Modulating the immune response using antibody-drug conjugates

ABSTRACT

The invention provides methods and compositions for modulating the immune response in a subject, such as decreasing the activity of CD30 +  T regulatory cells and increasing the ratio of CD8 +  T cells to CD30 +  T regulatory cells, through administration of antibody drug-conjugates that bind to CD30. The invention also provides articles of manufacture or kits comprising said antibody drug-conjugates that bind to CD30 for modulating the immune response.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims priority to U.S. Provisional Application No.62/572,345 filed on Oct. 13, 2017, U.S. Provisional Application No.62/576,017 filed on Oct. 23, 2017, and U.S. Provisional Application No.62/657,511 filed on Apr. 13, 2018; the contents of each of which areincorporated herein by reference in their entirety.

SUBMISSION OF SEQUENCE LISTING ON ASCII TEXT FILE

The content of the following submission on ASCII text file isincorporated herein by reference in its entirety: a computer readableform (CRF) of the Sequence Listing (file name: 761682000140SEQLIST.TXT,date recorded: Oct. 8, 2018, size: 6 KB).

FIELD OF THE INVENTION

The present invention relates to anti-CD30 antibody-drug conjugates andmethods of using the same to modulate the immune response for thetreatment of cancer in a subject.

BACKGROUND OF THE INVENTION

CD30 is a 120 kilodalton membrane glycoprotein (Froese et al., 1987, J.Immunol. 139: 2081-87) and a member of the TNF-receptor superfamily thathas been shown to be a marker of malignant cells in Hodgkin's lymphomaand anaplastic large cell lymphoma (ALCL), a subset of non-Hodgkin'slymphoma (NHL) (DUrkop et al., 1992, Cell 88:421-427). CD30 has beenfound to be highly expressed on the cell surface of all Hodgkin'slymphomas and the majority of ALCL (Josimovic-Alasevic et al., 1989,Eur. J. Immunol. 19:157-162).

CD30 was originally identified by the monoclonal antibody Ki-1 (Schwabet al., 1982, Nature 299:65-67). This monoclonal antibody was developedagainst Hodgkin and Reed-Sternberg (H-RS) cells, the malignant cells ofHodgkin's lymphoma. A second monoclonal antibody, capable of binding aformalin resistant epitope different from that recognized by Ki-1, wassubsequently described (Schwarting et al, 1989 Blood 74:1678-1689). Theidentification of four additional antibodies resulted in the creation ofthe CD30 cluster at the Third Leucocyte Typing Workshop in 1986(McMichael, A, ed., 1987, Leukocyte Typing 111 (Oxford: OxfordUniversity Press)). Monoclonal antibodies specific for the CD30 antigenhave been explored as vehicles for the delivery of cytostatic drugs,plant toxins and radioisotopes to cancerous cells expressing CD30 inboth preclinical models and clinical studies (Engert et al., 1990,Cancer Research 50:84-88; Barth et al., 2000, Blood 95:3909-3914). Inpatients with Hodgkin's lymphoma, targeting of the CD30 antigen could beachieved with low doses of the anti-CD30 antibody, BerH2 (Falini et al.,1992, British Journal of Haematology 82:38-45). Yet, despite successfulin vivo targeting of the malignant tumor cells, none of the patientsexperienced tumor regression. In a subsequent clinical trial, the toxinsaporin was chemically conjugated to the BerH2 antibody and all fourpatients demonstrated rapid and substantial reductions in tumor mass(Falini et al., 1992, Lancet 339:1195-1196). However, in vitro studiesusing an antibody drug-conjugate (ADC) where the toxin dgA wasconjugated to the Ki-1 antibody demonstrated only moderate efficacy whenadministered to patients with resistant HL in a Phase 1 clinical trial(Schnell et al., 2002, Clinical Cancer Research, 8(6):1779-1786).

T regulatory cells (Tregs) are essential modulators of T cell immuneresponses, limiting chronic inflammation and protecting normal tissuesfrom autoimmunity. T regulatory cells are also implicated in maintainingimmune-suppressive conditions in the tumor microenvironment, abrogatingcytotoxic anti-tumor immunosurveillance. Analysis of clinical tumorsamples has shown increased densities of intratumoral Tregs associatedwith poor clinical outcomes in a number of cancer types (Fridman, 2012,Nature Reviews Cancer; Charoentong, 2017, Cell Reports 18: 248-262).Recent transcriptomic analyses of intratumoral Tregs isolated frombreast, lung, and colorectal cancer tissues showed TNFSFR8 (CD30) to beamong transcripts differentially upregulated compared to Tregs isolatedfrom adjacent normal tissue and circulating in blood (Plitas, 2016,Immunity, 45: 1122-1134; De Simone, 2016, Immunity, 45: 1135-1147). Thefunctional significance of heightened CD30 transcript expression inTregs remains unclear. Given the protective role of Tregs in promotingimmune homeostasis in normal tissues, there is considerable interest indeveloping cancer therapeutics that preferentially target intratumoralTregs, while sparing those in non-diseased tissues. Therefore, thereappears to be a need for therapies that can selectively control theactivity of immune cells that are involved in pathogenesis of cancer,such as the activity of T regulatory cells.

All references cited herein, including patent applications, patentpublications, and scientific literature, are herein incorporated byreference in their entirety, as if each individual reference werespecifically and individually indicated to be incorporated by reference.

SUMMARY

In one aspect, the present invention provides for a method of decreasingthe activity of CD30⁺ T regulatory (Treg) cells in a subject havingcancer comprising administering to the subject an antibody-drugconjugate, wherein the antibody-drug conjugate comprises an anti-CD30antibody or an antigen-binding portion thereof conjugated to amonomethyl auristatin.

In some embodiments, decreasing the activity of CD30⁺ Treg cellscomprises a decrease in the number of CD30⁺ Treg cells. In someembodiments, the number of CD30⁺ Treg cells is decreased relative to thenumber of one or more other types of CD4⁺ T cells. In some embodiments,the one or more other types of CD4⁺ T cells comprise Th1 cells, Th2cells or Th17 cells. In some embodiments, the one or more other types ofCD4⁺ T cells comprise Th1 CD30⁺ cells, Th2 CD30⁺ cells or Th17 CD30⁺cells. In some embodiments, the number of CD30⁺ Treg cells is decreasedrelative to the number of CD30⁺ Treg cells in the subject prior toadministration of the antibody-drug conjugate.

In some embodiments, decreasing the activity of CD30⁺ Treg cellscomprises a decrease in the function of CD30⁺ Treg cells. In someembodiments, the decrease in the function of CD30⁺ Treg cells isrelative to the function of CD30⁺ Treg cells in a subject prior toadministration of the antibody-drug conjugate.

In some embodiments, the CD30⁺ Treg cells are CD30⁺ inducible Tregulatory (iTreg) cells or CD30⁺ peripheral T regulatory (pTreg) cells.

In some embodiments, the monomethyl auristatin is monomethyl auristatinE (MMAE). In some embodiments, the monomethyl auristatin is monomethylauristatin F (MMAF).

In some embodiments, the anti-CD30 antibody is monoclonal anti-CD30antibody AC10. In some embodiments, the anti-CD30 antibody is cAC10. Insome embodiments, the antibody-drug conjugate is brentuximab vedotin.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region and a light chain variable region, wherein the heavychain variable region comprises:

-   -   (i) a CDR-H1 comprising the amino acid sequence of SEQ ID NO: 1;    -   (ii) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:        2; and    -   (iii) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:        3; and        wherein the light chain variable region comprises:    -   (i) a CDR-L1 comprising the amino acid sequence of SEQ ID NO: 4;    -   (ii) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:        5; and    -   (iii) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:        6.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region comprising the amino acid sequence of SEQ ID NO: 7 and alight chain variable region comprising the amino acid sequence of SEQ IDNO: 8.

In some embodiments, the antibody-drug conjugate comprises a linkerbetween the anti-CD30 antibody or antigen-binding portion thereof andthe monomethyl auristatin. In some embodiments, the linker is selectedfrom the group consisting of a cleavable linker and a non-cleavablelinker. In some embodiments, the linker is a cleavable peptide linker.In some embodiments, the linker is a protease-cleavable linker. In someembodiments, the protease cleavable linker is comprises a thiolreactivespacer and a dipeptide. In some embodiments, the protease cleavablelinker comprises a thiolreactive maleimidocaproyl spacer, avaline-citrulline dipeptide, and a p-amino-benzyloxycarbonyl spacer. Insome embodiments, the cleavable peptide linker has a formula:-MC-vc-PAB-. In some embodiments, the linker is a non-cleavable linkerhaving a formula: -MC-.

In some embodiments, the subject has been previously treated for thecancer. In some embodiments, the subject did not respond to treatment orrelapsed after first-line treatment. In some embodiments, the subjecthas not previously been treated for the cancer.

In some embodiments, the cancer is a lymphoma. In some embodiments, thelymphoma is a T-cell lymphoma. In some embodiments, the lymphoma is aB-cell lymphoma.

In some embodiments, the lymphoma is a non-Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the non-Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject has not beenpreviously treated for the non-Hodgkin lymphoma. In some embodiments,the non-Hodgkin lymphoma is a mature T-cell lymphoma. In someembodiments, the non-Hodgkin lymphoma is diffuse large B-cell lymphoma(DLBCL), peripheral T-cell lymphoma (PTCL), anaplastic large celllymphoma (ALCL) or cutaneous T-cell lymphoma (CTCL). In someembodiments, the non-Hodgkin lymphoma is cutaneous T-cell lymphoma(CTCL). In some embodiments, the cutaneous T-cell lymphoma (CTCL) is amycosis fungoides (MF). In some embodiments, the mycosis fungoides is aCD30-positive mycosis fungoides (MF). In some embodiments, the cutaneousT-cell lymphoma (CTCL) is a primary cutaneous anaplastic large celllymphoma (pcALCL). In some embodiments, the subject has received priorsystemic treatment. In some embodiments, the non-Hodgkin lymphoma isanaplastic large cell lymphoma (ALCL). In some embodiments, theanaplastic large cell lymphoma (ALCL) is a systemic anaplastic largecell lymphoma (sALCL).

In some embodiments, the lymphoma is a Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject relapsed afterautologous stem cell transplant. In some embodiments, the subjectrelapsed after first-line treatment and the subject is ineligible forautologous stem cell transplant. In some embodiments, the subject hasnot been previously treated for the Hodgkin lymphoma. In someembodiments, the Hodgkin lymphoma is classical Hodgkin lymphoma (cHL).In some embodiments, the classical Hodgkin lymphoma (cHL) is advancedcHL. In some embodiments, the subject has been previously treated forcHL. In some embodiments, the subject has not been previously treatedfor cHL.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents capable of modulating the immuneresponse. In some embodiments, the one or more additional therapeuticagents is not an antibody or antigen-binding fragment thereof. In someembodiments, the one or more additional therapeutic agents is anantibody or antigen-binding fragment thereof.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents. In some embodiments, the one or moreadditional therapeutic agents is a chemotherapy regimen consistingessentially of doxorubicin, vinblastine, and dacarbazine (AVD). In someembodiments, the one or more additional therapeutic agents is achemotherapy regimen consisting essentially of Cyclophosphamide,Doxorubicin, and Prednisone (CHP). In some embodiments, the one or moreadditional therapeutic agents is an alkylating agent, an anthracycline,an antibiotic, an antifolate, an antimetabolite, an antitubulin agent,an auristatin, a chemotherapy sensitizer, a DNA minor groove binder, aDNA replication inhibitor, a duocarmycin, an etoposide, a fluorinatedpyrimidine, a lexitropsin, a nitrosourea, a platinol, a purineantimetabolite, a puromycin, a radiation sensitizer, a steroid, ataxane, a topoisomerase inhibitor, and/or a vinca alkaloid. In someembodiments, the one or more additional therapeutic agents is selectedfrom the group consisting of adriamycin, an androgen, anthramycin (AMC),asparaginase, 5-azacytidine, azathioprine, bleomycin, busulfan,buthionine sulfoximine, camptothecin, carboplatin, carmustine (BSNU),CC-1065, chlorambucil, cisplatin, colchicine, cyclophosphamide,cytarabine, cytidine arabinoside, cytochalasin B, dacarbazine,dactinomycin (formerly actinomycin), daunorubicin, decarbazine,docetaxel, doxorubicin, an estrogen, 5-fluordeoxyuridine,5-fluorouracil, gramicidin D, hydroxydaunorubicin, hydroxyurea,idarubicin, ifosfamide, irinotecan, lomustine (CCNU), mechlorethamine,melphalan, 6-mercaptopurine, methotrexate, mithramycin, mitomycin C,mitoxantrone, nitroimidazole, paclitaxel, plicamycin, prednisone,prednisolone, procarbizine, streptozotocin, tenoposide, 6-thioguanine,thioTEPA, topotecan, vinblastine, vincristine, vinorelbine, VP-16 andVM-26. In some embodiments, the one or more additional therapeuticagents is an antibody or antigen-binding fragment thereof.

In some embodiments, the subject has cHL that has not been previouslytreated and the one or more additional therapeutic agents areadriamycin, dacarabazine and vinblastine (AVD). In some embodiments, thecHL is advanced cHL.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and the one or more additional therapeuticagents are cyclophosphamide, hydroxydaunorubicin and prednisone (CHP).In some embodiments, the subject has cutaneous T-cell lymphoma (CTCL)and has been previously treated.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and the one or more additional therapeuticagents are cyclophosphamide, hydroxydaunorubicin and prednisolone.

In some embodiments, the method further comprises treating the subjectwith irradiation.

In another aspect, the present invention provides for a method ofincreasing the ratio of CD8⁺ T cells to CD30⁺ T regulatory (Treg) cellsin a subject having cancer comprising administering to the subject anantibody-drug conjugate, wherein the antibody-drug conjugate comprisesan anti-CD30 antibody or an antigen-binding portion thereof conjugatedto a monomethyl auristatin. In some embodiments, ratio of CD8⁺ T cellsto CD30⁺ Treg cells is increased relative to the ratio of CD8⁺ T cellsto CD30⁺ Treg cells in the subject prior to the administration of theantibody-drug conjugate.

In some embodiments, the CD30⁺ Treg cells are CD30⁺ inducible Tregulatory (iTreg) cells or CD30⁺ peripheral T regulatory (pTreg) cells.

In some embodiments, the monomethyl auristatin is monomethyl auristatinE (MMAE). In some embodiments, the monomethyl auristatin is monomethylauristatin F (MMAF).

In some embodiments, the anti-CD30 antibody is monoclonal anti-CD30antibody AC10. In some embodiments, the anti-CD30 antibody is cAC10. Insome embodiments, the antibody-drug conjugate is brentuximab vedotin.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region and a light chain variable region, wherein the heavychain variable region comprises:

-   -   (i) a CDR-H1 comprising the amino acid sequence of SEQ ID NO: 1;    -   (ii) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:        2; and    -   (iii) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:        3; and        wherein the light chain variable region comprises:    -   (i) a CDR-L1 comprising the amino acid sequence of SEQ ID NO: 4;    -   (ii) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:        5; and    -   (iii) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:        6.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region comprising the amino acid sequence of SEQ ID NO: 7 and alight chain variable region comprising the amino acid sequence of SEQ IDNO: 8.

In some embodiments, the antibody-drug conjugate comprises a linkerbetween the anti-CD30 antibody or antigen-binding portion thereof andthe monomethyl auristatin. In some embodiments, the linker is selectedfrom the group consisting of a cleavable linker and a non-cleavablelinker. In some embodiments, the linker is a cleavable peptide linker.In some embodiments, the linker is a protease-cleavable linker. In someembodiments, the protease cleavable linker is comprises a thiolreactivespacer and a dipeptide. In some embodiments, the protease cleavablelinker comprises a thiolreactive maleimidocaproyl spacer, avaline-citrulline dipeptide, and a p-amino-benzyloxycarbonyl spacer. Insome embodiments, the cleavable peptide linker has a formula:-MC-vc-PAB-. In some embodiments, the linker is a non-cleavable linkerhaving a formula: -MC-.

In some embodiments, the subject has been previously treated for thecancer. In some embodiments, the subject did not respond to treatment orrelapsed after first-line treatment. In some embodiments, the subjecthas not previously been treated for the cancer.

In some embodiments, the cancer is a lymphoma. In some embodiments, thelymphoma is a T-cell lymphoma. In some embodiments, the lymphoma is aB-cell lymphoma.

In some embodiments, the lymphoma is a non-Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the non-Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject has not beenpreviously treated for the non-Hodgkin lymphoma. In some embodiments,the non-Hodgkin lymphoma is a mature T-cell lymphoma. In someembodiments, the non-Hodgkin lymphoma is diffuse large B-cell lymphoma(DLBCL), peripheral T-cell lymphoma (PTCL), anaplastic large celllymphoma (ALCL) or cutaneous T-cell lymphoma (CTCL). In someembodiments, the non-Hodgkin lymphoma is cutaneous T-cell lymphoma(CTCL). In some embodiments, the cutaneous T-cell lymphoma (CTCL) is amycosis fungoides (MF). In some embodiments, the mycosis fungoides is aCD30-positive mycosis fungoides (MF). In some embodiments, the cutaneousT-cell lymphoma (CTCL) is a primary cutaneous anaplastic large celllymphoma (pcALCL). In some embodiments, the subject has received priorsystemic treatment. In some embodiments, the non-Hodgkin lymphoma isanaplastic large cell lymphoma (ALCL). In some embodiments, theanaplastic large cell lymphoma (ALCL) is a systemic anaplastic largecell lymphoma (sALCL).

In some embodiments, the lymphoma is a Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject relapsed afterautologous stem cell transplant. In some embodiments, the subjectrelapsed after first-line treatment and the subject is ineligible forautologous stem cell transplant. In some embodiments, the subject hasnot been previously treated for the Hodgkin lymphoma. In someembodiments, the Hodgkin lymphoma is classical Hodgkin lymphoma (cHL).In some embodiments, the classical Hodgkin lymphoma (cHL) is advancedcHL. In some embodiments, the subject has been previously treated forcHL. In some embodiments, the subject has not been previously treatedfor cHL.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents capable of modulating the immuneresponse. In some embodiments, the one or more additional therapeuticagents is not an antibody or antigen-binding fragment thereof. In someembodiments, the one or more additional therapeutic agents is anantibody or antigen-binding fragment thereof.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents. In some embodiments, the one or moreadditional therapeutic agents is a chemotherapy regimen consistingessentially of doxorubicin, vinblastine, and dacarbazine (AVD). In someembodiments, the one or more additional therapeutic agents is achemotherapy regimen consisting essentially of Cyclophosphamide,Doxorubicin, and Prednisone (CHP). In some embodiments, the one or moreadditional therapeutic agents is an alkylating agent, an anthracycline,an antibiotic, an antifolate, an antimetabolite, an antitubulin agent,an auristatin, a chemotherapy sensitizer, a DNA minor groove binder, aDNA replication inhibitor, a duocarmycin, an etoposide, a fluorinatedpyrimidine, a lexitropsin, a nitrosourea, a platinol, a purineantimetabolite, a puromycin, a radiation sensitizer, a steroid, ataxane, a topoisomerase inhibitor, and/or a vinca alkaloid. In someembodiments, the one or more additional therapeutic agents is selectedfrom the group consisting of adriamycin, an androgen, anthramycin (AMC),asparaginase, 5-azacytidine, azathioprine, bleomycin, busulfan,buthionine sulfoximine, camptothecin, carboplatin, carmustine (BSNU),CC-1065, chlorambucil, cisplatin, colchicine, cyclophosphamide,cytarabine, cytidine arabinoside, cytochalasin B, dacarbazine,dactinomycin (formerly actinomycin), daunorubicin, decarbazine,docetaxel, doxorubicin, an estrogen, 5-fluordeoxyuridine,5-fluorouracil, gramicidin D, hydroxydaunorubicin, hydroxyurea,idarubicin, ifosfamide, irinotecan, lomustine (CCNU), mechlorethamine,melphalan, 6-mercaptopurine, methotrexate, mithramycin, mitomycin C,mitoxantrone, nitroimidazole, paclitaxel, plicamycin, prednisone,prednisolone, procarbizine, streptozotocin, tenoposide, 6-thioguanine,thioTEPA, topotecan, vinblastine, vincristine, vinorelbine, VP-16 andVM-26. In some embodiments, the one or more additional therapeuticagents is an antibody or antigen-binding fragment thereof.

In some embodiments, the subject has cHL that has not been previouslytreated and the one or more additional therapeutic agents areadriamycin, dacarabazine and vinblastine. In some embodiments, the cHLis advanced cHL.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and the one or more additional therapeuticagents are cyclophosphamide, hydroxydaunorubicin and prednisone.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and the one or more additional therapeuticagents are cyclophosphamide, hydroxydaunorubicin and prednisolone.

In some embodiments, the method further comprises treating the subjectwith irradiation.

In another aspect, the present invention provides for a method ofmodulating the immune response in a subject having cancer comprisingadministering to the subject an antibody-drug conjugate, wherein theantibody-drug conjugate comprises an anti-CD30 antibody or anantigen-binding portion thereof conjugated to a monomethyl auristatin,wherein the modulation comprises increasing the ratio of CD8⁺ T cells toCD30⁺ T regulatory (Treg) cells in the subject. In some embodiments,ratio of CD8⁺ T cells to CD30⁺ Treg cells is increased relative to theratio of CD8⁺ T cells to CD30⁺ Treg cells in the subject prior to theadministration of the antibody-drug conjugate.

In some embodiments, the CD30⁺ Treg cells are CD30⁺ inducible Tregulatory (iTreg) cells or CD30⁺ peripheral T regulatory (pTreg) cells.

In some embodiments, the monomethyl auristatin is monomethyl auristatinE (MMAE). In some embodiments, the monomethyl auristatin is monomethylauristatin F (MMAF).

In some embodiments, the anti-CD30 antibody is monoclonal anti-CD30antibody AC10. In some embodiments, the anti-CD30 antibody is cAC10. Insome embodiments, the antibody-drug conjugate is brentuximab vedotin.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region and a light chain variable region, wherein the heavychain variable region comprises:

-   -   (i) a CDR-H1 comprising the amino acid sequence of SEQ ID NO: 1;    -   (ii) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:        2; and    -   (iii) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:        3; and        wherein the light chain variable region comprises:    -   (i) a CDR-L1 comprising the amino acid sequence of SEQ ID NO: 4;    -   (ii) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:        5; and    -   (iii) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:        6.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region comprising the amino acid sequence of SEQ ID NO: 7 and alight chain variable region comprising the amino acid sequence of SEQ IDNO: 8.

In some embodiments, the antibody-drug conjugate comprises a linkerbetween the anti-CD30 antibody or antigen-binding portion thereof andthe monomethyl auristatin. In some embodiments, the linker is selectedfrom the group consisting of a cleavable linker and a non-cleavablelinker. In some embodiments, the linker is a cleavable peptide linker.In some embodiments, the linker is a protease-cleavable linker. In someembodiments, the protease cleavable linker is comprises a thiolreactivespacer and a dipeptide. In some embodiments, the protease cleavablelinker comprises a thiolreactive maleimidocaproyl spacer, avaline-citrulline dipeptide, and a p-amino-benzyloxycarbonyl spacer. Insome embodiments, the cleavable peptide linker has a formula:-MC-vc-PAB-. In some embodiments, the linker is a non-cleavable linkerhaving a formula: -MC-.

In some embodiments, the subject has been previously treated for thecancer. In some embodiments, the subject did not respond to treatment orrelapsed after first-line treatment. In some embodiments, the subjecthas not previously been treated for the cancer.

In some embodiments, the cancer is a lymphoma. In some embodiments, thelymphoma is a T-cell lymphoma. In some embodiments, the lymphoma is aB-cell lymphoma.

In some embodiments, the lymphoma is a non-Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the non-Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject has not beenpreviously treated for the non-Hodgkin lymphoma. In some embodiments,the non-Hodgkin lymphoma is a mature T-cell lymphoma. In someembodiments, the non-Hodgkin lymphoma is diffuse large B-cell lymphoma(DLBCL), peripheral T-cell lymphoma (PTCL), anaplastic large celllymphoma (ALCL) or cutaneous T-cell lymphoma (CTCL). In someembodiments, the non-Hodgkin lymphoma is cutaneous T-cell lymphoma(CTCL). In some embodiments, the cutaneous T-cell lymphoma (CTCL) is amycosis fungoides (MF). In some embodiments, the mycosis fungoides is aCD30-positive mycosis fungoides (MF). In some embodiments, the cutaneousT-cell lymphoma (CTCL) is a primary cutaneous anaplastic large celllymphoma (pcALCL). In some embodiments, the subject has received priorsystemic treatment. In some embodiments, the non-Hodgkin lymphoma isanaplastic large cell lymphoma (ALCL). In some embodiments, theanaplastic large cell lymphoma (ALCL) is a systemic anaplastic largecell lymphoma (sALCL).

In some embodiments, the lymphoma is a Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject relapsed afterautologous stem cell transplant. In some embodiments, the subjectrelapsed after first-line treatment and the subject is ineligible forautologous stem cell transplant. In some embodiments, the subject hasnot been previously treated for the Hodgkin lymphoma. In someembodiments, the Hodgkin lymphoma is classical Hodgkin lymphoma (cHL).In some embodiments, the classical Hodgkin lymphoma (cHL) is advancedcHL. In some embodiments, the subject has been previously treated forcHL. In some embodiments, the subject has not been previously treatedfor cHL.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents capable of modulating the immuneresponse. In some embodiments, the one or more additional therapeuticagents is not an antibody or antigen-binding fragment thereof. In someembodiments, the one or more additional therapeutic agents is anantibody or antigen-binding fragment thereof.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents. In some embodiments, the one or moreadditional therapeutic agents is a chemotherapy regimen consistingessentially of doxorubicin, vinblastine, and dacarbazine (AVD). In someembodiments, the one or more additional therapeutic agents is achemotherapy regimen consisting essentially of Cyclophosphamide,Doxorubicin, and Prednisone (CHP). In some embodiments, the one or moreadditional therapeutic agents is an alkylating agent, an anthracycline,an antibiotic, an antifolate, an antimetabolite, an antitubulin agent,an auristatin, a chemotherapy sensitizer, a DNA minor groove binder, aDNA replication inhibitor, a duocarmycin, an etoposide, a fluorinatedpyrimidine, a lexitropsin, a nitrosourea, a platinol, a purineantimetabolite, a puromycin, a radiation sensitizer, a steroid, ataxane, a topoisomerase inhibitor, and/or a vinca alkaloid. In someembodiments, the one or more additional therapeutic agents is selectedfrom the group consisting of adriamycin, an androgen, anthramycin (AMC),asparaginase, 5-azacytidine, azathioprine, bleomycin, busulfan,buthionine sulfoximine, camptothecin, carboplatin, carmustine (BSNU),CC-1065, chlorambucil, cisplatin, colchicine, cyclophosphamide,cytarabine, cytidine arabinoside, cytochalasin B, dacarbazine,dactinomycin (formerly actinomycin), daunorubicin, decarbazine,docetaxel, doxorubicin, an estrogen, 5-fluordeoxyuridine,5-fluorouracil, gramicidin D, hydroxydaunorubicin, hydroxyurea,idarubicin, ifosfamide, irinotecan, lomustine (CCNU), mechlorethamine,melphalan, 6-mercaptopurine, methotrexate, mithramycin, mitomycin C,mitoxantrone, nitroimidazole, paclitaxel, plicamycin, prednisone,prednisolone, procarbizine, streptozotocin, tenoposide, 6-thioguanine,thioTEPA, topotecan, vinblastine, vincristine, vinorelbine, VP-16 andVM-26. In some embodiments, the one or more additional therapeuticagents is an antibody or antigen-binding fragment thereof.

In some embodiments, the subject has cHL that has not been previouslytreated and the one or more additional therapeutic agents areadriamycin, dacarabazine and vinblastine. In some embodiments, the cHLis advanced cHL.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and the one or more additional therapeuticagents are cyclophosphamide, hydroxydaunorubicin and prednisone.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and the one or more additional therapeuticagents are cyclophosphamide, hydroxydaunorubicin and prednisolone.

In some embodiments, the method further comprises treating the subjectwith irradiation.

It is to be understood that one, some, or all of the properties of thevarious embodiments described herein may be combined to form otherembodiments of the present invention. These and other aspects of theinvention will become apparent to one of skill in the art. These andother embodiments of the invention are further described by the detaileddescription that follows.

BRIEF DESCRIPTION OF THE DRAWINGS

FIGS. 1A and 1B is a series of graphs showing that brentuximab vedotin(BV) impaired T regulatory cells in vitro. A) BV drove a dose-dependentreduction in total viable iTreg numbers from five separate donors. B) BVshowed enhanced depletion of CD30⁺iTregs. Cell counts are shown as thepercent of untreated control. ADC indicates antibody drug-conjugate. IgGMMAE indicates control ADC.

FIG. 2A-D is a series of graphs showing that treatment with BV reducedthe number of inducible T regulatory cells while relieving therepression of in vitro CD8⁺ T cell proliferation. Increasing the iTregto CD8⁺ T cell ratio (iTreg:CD8 ratio) of cells isolated from A) Donor 1or B) Donor 2 abrogated T cell expansion. Increasing concentrations ofBV treatment selectively reduced iTregs and augmented CD8⁺ T cellaccumulation for both C) Donor 1 and D) Donor 2. Cell counts are shownas the percent of untreated control. ADC indicates antibodydrug-conjugate. IgG MMAE indicates control ADC.

FIGS. 3A and 3B is a series of graphs showing that BV depleted naturallyoccurring CD30⁺ blood Tregs but not CD30⁺ CD8⁺ T cells in vitro. A) BVdrove a dose-dependent reduction of viable CD30⁺ Treg numbers from fourseparate donors. B) BV did not deplete CD30⁺ CD8⁺ T cells. ADC indicatesantibody drug-conjugate. IgG MMAE indicates control ADC.

FIG. 4A-C is a series of graphs showing that BV reduced T regulatorycells and increased the CD8⁺ T cell to Treg ratio in a xeno-GVHD mousemodel. A) BV significantly reduced human T regulatory cells in thespleen compared to PBS alone (untreated). B) Splenic CD8⁺ T cells wereunaffected by BV treatment with a trend toward increased numbers. C) BVtreatment increased the CD8⁺ T cell/Treg ratio in vivo.

FIG. 5 is a graph showing that single treatment with BV in patients withclassical Hodgkin lymphoma resulted in the reduction of T helper cellssubset populations.

FIG. 6 is a graph showing the expression of CD30 in T cell subtypesisolated from human blood.

FIG. 7 is a graph showing that single treatment with BV in patients withclassical Hodgkin lymphoma reduced the number of T regulatory cells thatexpressed CD30 (CD30+) as compared to the number of T regulatory cellsthat did not express CD30 (CD30−). BSLN indicates baseline measurement.

FIGS. 8A and 8B is a series of graphs showing that CD30 expression isenriched on CD25^(hi) CCR4^(hi) FoxP3^(hi) effector Tregs in PBMC. A)CD30 is most frequently expressed by T regulatory cells compared to CD4⁺and CD8⁺ memory and naïve T cell subsets. B) Expression of CD30 ishighly associated with the effector T regulatory subset (FoxP3^(hi)CD25^(hi) CCR4^(hi)).

FIG. 9A-D is a series of graphs showing that activated T regulatorycells demonstrate heightened CD30 receptor expression and payloaddelivery, along with impaired drug efflux capacity. A) A higherproportion of enriched T regulatory cells express CD30 compared to CD4⁺and CD8⁺ T cells following activation. B) Enriched T regulatory cellshave an increased magnitude of expression of CD30 by Mean FluorescenceIntensity (MFI) compared to CD4⁺ and CD8⁺ T cells following activation.C) T regulatory cells show accelerated and increased release offluorescent payload from a conditionally fluorescent anti-CD30 mAbrelative to CD4⁺ and CD8⁺ T cells in an internalization assay. D) Tregulatory cells show the slowest rhodamine-123 efflux among T cellsubsets while CD8⁺ T cells show rapid clearance of intracellularrhodamine-123 in a rhodamine 123 efflux assay.

DETAILED DESCRIPTION I. Definitions

In order that the present disclosure can be more readily understood,certain terms are first defined. As used in this application, except asotherwise expressly provided herein, each of the following terms shallhave the meaning set forth below. Additional definitions are set forththroughout the application.

The term “and/or” where used herein is to be taken as specificdisclosure of each of the two specified features or components with orwithout the other. Thus, the term “and/or” as used in a phrase such as“A and/or B” herein is intended to include “A and B,” “A or B,” “A”(alone), and “B” (alone). Likewise, the term “and/or” as used in aphrase such as “A, B, and/or C” is intended to encompass each of thefollowing aspects: A, B, and C; A, B, or C; A or C; A or B; B or C; Aand C; A and B; B and C; A (alone); B (alone); and C (alone).

The term “about” as used herein refers to the usual error range for therespective value readily known to the skilled person in this technicalfield. Reference to “about” a value or parameter herein includes (anddescribes) embodiments that are directed to that value or parameter perse.

It is understood that aspects and embodiments of the invention describedherein include “comprising,” “consisting,” and “consisting essentiallyof” aspects and embodiments.

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this disclosure is related. For example, the ConciseDictionary of Biomedicine and Molecular Biology, Juo, Pei-Show, 2nd ed.,2002, CRC Press; The Dictionary of Cell and Molecular Biology, 3rd ed.,1999, Academic Press; and the Oxford Dictionary Of Biochemistry AndMolecular Biology, Revised, 2000, Oxford University Press, provide oneof skill with a general dictionary of many of the terms used in thisdisclosure.

Units, prefixes, and symbols are denoted in their Systeme Internationalde Unites (SI) accepted form. Numeric ranges are inclusive of thenumbers defining the range. The headings provided herein are notlimitations of the various aspects of the disclosure, which can be hadby reference to the specification as a whole. Accordingly, the termsdefined immediately below are more fully defined by reference to thespecification in its entirety.

“Administering” refers to the physical introduction of a therapeuticagent to a subject, using any of the various methods and deliverysystems known to those skilled in the art. Exemplary routes ofadministration include intravenous, intramuscular, subcutaneous,intraperitoneal, spinal or other parenteral routes of administration,for example by injection or infusion. The phrase “parenteraladministration” as used herein means modes of administration other thanenteral and topical administration, usually by injection, and includes,without limitation, intravenous, intramuscular, intraarterial,intrathecal, intralymphatic, intralesional, intracapsular, intraorbital,intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous,subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal,epidural and intrasternal injection and infusion, as well as in vivoelectroporation. A therapeutic agent can be administered via anon-parenteral route, or orally. Other non-parenteral routes include atopical, epidermal or mucosal route of administration, for example,intranasally, vaginally, rectally, sublingually or topically.Administering can also be performed, for example, once, a plurality oftimes, and/or over one or more extended periods.

An “adverse event” (AE) as used herein is any unfavorable and generallyunintended or undesirable sign (including an abnormal laboratoryfinding), symptom, or disease associated with the use of a medicaltreatment. A medical treatment can have one or more associated AEs andeach AE can have the same or different level of severity. Reference tomethods capable of “altering adverse events” means a treatment regimethat decreases the incidence and/or severity of one or more AEsassociated with the use of a different treatment regime.

An “antibody” (Ab) shall include, without limitation, a glycoproteinimmunoglobulin which binds specifically to an antigen and comprises atleast two heavy (H) chains and two light (L) chains interconnected bydisulfide bonds, or an antigen-binding portion thereof. Each H chaincomprises a heavy chain variable region (abbreviated herein as V_(H))and a heavy chain constant region. The heavy chain constant regioncomprises at least three constant domains, C_(H1), C_(H2) and C_(H3).Each light chain comprises a light chain variable region (abbreviatedherein as V_(L)) and a light chain constant region. The light chainconstant region comprises one constant domain, C_(L). The V_(H) andV_(L) regions can be further subdivided into regions ofhypervariability, termed complementarity determining regions (CDRs),interspersed with regions that are more conserved, termed frameworkregions (FRs). Each V_(H) and V_(L) comprises three CDRs and four FRs,arranged from amino-terminus to carboxy-terminus in the following order:FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4. The variable regions of theheavy and light chains contain a binding domain that interacts with anantigen. The constant regions of the antibodies can mediate the bindingof the immunoglobulin to host tissues or factors, including variouscells of the immune system (e.g., effector cells) and the firstcomponent (C1q) of the classical complement system.

An immunoglobulin can derive from any of the commonly known isotypes,including but not limited to IgA, secretory IgA, IgG, and IgM. IgGsubclasses are also well known to those in the art and include but arenot limited to human IgG1, IgG2, IgG3 and IgG4. “Isotype” refers to theantibody class or subclass (e.g., IgM or IgG1) that is encoded by theheavy chain constant region genes. The term “antibody” includes, by wayof example, both naturally occurring and non-naturally occurringantibodies; monoclonal and polyclonal antibodies; chimeric and humanizedantibodies; human or non-human antibodies; wholly synthetic antibodies;and single chain antibodies. A non-human antibody can be humanized byrecombinant methods to reduce its immunogenicity in man. Where notexpressly stated, and unless the context indicates otherwise, the term“antibody” also includes an antigen-binding fragment or anantigen-binding portion of any of the aforementioned immunoglobulins,and includes a monovalent and a divalent fragment or portion, and asingle chain antibody.

An “isolated antibody” refers to an antibody that is substantially freeof other antibodies having different antigenic specificities (e.g., anisolated antibody that binds specifically to CD30 is substantially freeof antibodies that bind specifically to antigens other than CD30). Anisolated antibody that binds specifically to CD30 can, however, havecross-reactivity to other antigens, such as CD30 molecules fromdifferent species. Moreover, an isolated antibody can be substantiallyfree of other cellular material and/or chemicals. In one embodiment, anantibody includes a conjugate attached to another agent (e.g., smallmolecule drug). In some embodiments, an anti-CD30 antibody includes aconjugate of an anti-CD30 antibody with a small molecule drug (e.g.,MMAE or MMAF).

The term “monoclonal antibody” (mAb) refers to a non-naturally occurringpreparation of antibody molecules of single molecular composition, i.e.,antibody molecules whose primary sequences are essentially identical,and which exhibits a single binding specificity and affinity for aparticular epitope. A monoclonal antibody is an example of an isolatedantibody. Monoclonal antibodies can be produced by hybridoma,recombinant, transgenic, or other techniques known to those skilled inthe art.

A “human antibody” (HuMAb) refers to an antibody having variable regionsin which both the FRs and CDRs are derived from human germlineimmunoglobulin sequences. Furthermore, if the antibody contains aconstant region, the constant region also is derived from human germlineimmunoglobulin sequences. The human antibodies of the disclosure caninclude amino acid residues not encoded by human germline immunoglobulinsequences (e.g., mutations introduced by random or site-specificmutagenesis in vitro or by somatic mutation in vivo). However, the term“human antibody,” as used herein, is not intended to include antibodiesin which CDR sequences derived from the germline of another mammalianspecies, such as a mouse, have been grafted onto human frameworksequences. The terms “human antibodies” and “fully human antibodies” andare used synonymously.

A “humanized antibody” refers to an antibody in which some, most, or allof the amino acids outside the CDRs of a non-human antibody are replacedwith corresponding amino acids derived from human immunoglobulins. Inone embodiment of a humanized form of an antibody, some, most, or all ofthe amino acids outside the CDRs have been replaced with amino acidsfrom human immunoglobulins, whereas some, most, or all amino acidswithin one or more CDRs are unchanged. Small additions, deletions,insertions, substitutions or modifications of amino acids arepermissible as long as they do not abrogate the ability of the antibodyto bind to a particular antigen. A “humanized antibody” retains anantigenic specificity similar to that of the original antibody. In someembodiments, the CDRs of a humanized antibody contain CDRs from anon-human, mammalian antibody. In other embodiments, the CDRs of ahumanized antibody contain CDRs from an engineered, synthetic antibody.

A “chimeric antibody” refers to an antibody in which the variableregions are derived from one species and the constant regions arederived from another species, such as an antibody in which the variableregions are derived from a mouse antibody and the constant regions arederived from a human antibody.

An “anti-antigen antibody” refers to an antibody that binds specificallyto the antigen. For example, an anti-CD30 antibody binds specifically toCD30.

An “antigen-binding portion” of an antibody (also called an“antigen-binding fragment”) refers to one or more fragments of anantibody that retain the ability to bind specifically to the antigenbound by the whole antibody. Examples of antibody fragments include butare not limited to Fv, Fab, Fab′, Fab′-SH, F(ab′)₂; diabodies; linearantibodies; single-chain antibody molecules (e.g. scFv); andmultispecific antibodies formed from antibody fragments. Papaindigestion of antibodies produces two identical antigen-bindingfragments, called “Fab” fragments, each with a single antigen-bindingsite, and a residual “Fc” fragment, whose name reflects its ability tocrystallize readily. Pepsin treatment yields an F(ab′)₂ fragment thathas two antigen-combining sites and is still capable of cross-linkingantigen.

The term “variable” refers to the fact that certain segments of thevariable domains differ extensively in sequence among antibodies. The Vdomain mediates antigen binding and defines the specificity of aparticular antibody for its particular antigen. However, the variabilityis not evenly distributed across the entire span of the variabledomains. Instead, it is concentrated in three segments calledcomplementarity determining regions (CDRs) both in the light-chain andthe heavy chain variable domains. The more highly conserved portions ofvariable domains are called the framework regions (FR). The variabledomains of native heavy and light chains each comprise four FR regions,largely adopting a beta-sheet configuration, connected by three CDRs,which form loops connecting, and in some cases forming part of, thebeta-sheet structure. The CDRs in each chain are held together in closeproximity by the FR regions and, with the CDRs from the other chain,contribute to the formation of the antigen binding site of antibodies(see Kabat et al, Sequences of Immunological Interest, Fifth Edition,National Institute of Health, Bethesda, Md. (1991)). The constantdomains are not involved directly in the binding of antibody to anantigen, but exhibit various effector functions, such as participationof the antibody in antibody-dependent cellular toxicity.

The “variable region” or “variable domain” of an antibody refers to thearnino-terminal domains of the heavy or light chain of the antibody. Thevariable domains of the heavy chain and light chain may be referred toas “VH’ and “VL”, respectively. These domains are generally the mostvariable parts of the antibody (relative to other antibodies of the sameclass) and contain the antigen binding sites.

The term “hypervariable region,” “HVR,” or “HV,” when used herein refersto the regions of an antibody-variable domain that are hypervariable insequence and/or form structurally defined loops. Generally, antibodiescomprise six HVRs; three in the VH (H1, H2, H3), and three in the VL(L1, L2, L3). In native antibodies, H3 and L3 display the most diversityof the six HVRs, and H3 in particular is believed to play a unique rolein conferring fine specificity to antibodies. See, e.g., Xu et al.Immunity 13:37-45 (2000); Johnson and Wu in Methods in Molecular Biology248:1-25 (Lo, ed., Human Press, Totowa, N.J., 2003)). Indeed, naturallyoccurring camelid antibodies consisting of a heavy chain only arefunctional and stable in the absence of light chain. See, e.g.,Hamers-Casterman et al., Nature 363:446-448 (1993) and Sheriff et al.,Nature Struct. Biol. 3:733-736 (1996).

A number of HVR delineations are in use and are encompassed herein. TheHVRs that are Kabat complementarity-determining regions (CDRs) are basedon sequence variability and are the most commonly used (Kabat et al.,Sequences of Proteins of Immunological Interest, 5^(th) Ed. PublicHealth Service, National Institute of Health, Bethesda, Md. (1991)).Chothia HVRs refer instead to the location of the structural loops(Chothia and Lesk J. Mol. Biol. 196:901-917 (1987)). The “contact” HVRsare based on an analysis of the available complex crystal structures.The residues from each of these HVRs are noted below.

Loop Kabat Chothia Contact L1 L24-L34 L26-L34 L30-L36 L2 L50-L56 L50-L56L46-L55 L3 L89-L97 L91-L96 L89-L96 H1 H31-H35B H26-H32 H30-H35B (KabatNumbering) H1 H31-H35 H26-H32 H30-H35 (Chothia Numbering) H2 H50-H65H53-H56 H47-H58 H3 H95-H102 H95-H102 H93-H101

Unless otherwise indicated, the variable-domain residues (HVR residuesand framework region residues) are numbered according to Kabat et al.,supra.

“Framework” or “FR” residues are those variable-domain residues otherthan the HVR residues as herein defined.

The expression “variable-domain residue-numbering as in Kabat” or“amino-acid-position numbering as in Kabat,” and variations thereof,refers to the numbering system used for heavy-chain variable domains orlight-chain variable domains of the compilation of antibodies in Kabatet al., supra. Using this numbering system, the actual linear amino acidsequence may contain fewer or additional amino acids corresponding to ashortening of, or insertion into, a FR or HVR of the variable domain.For example, a heavy-chain variable domain may include a single aminoacid insert (residue 52 a according to Kabat) after residue 52 of H2 andinserted residues (e.g. residues 82 a, 82 b, and 82 c, etc. according toKabat) after heavy-chain FR residue 82. The Kabat numbering of residuesmay be determined for a given antibody by alignment at regions ofhomology of the sequence of the antibody with a “standard” Kabatnumbered sequence.

As used herein, the term “specifically binds to” or is “specific for”refers to measurable and reproducible interactions such as bindingbetween a target and an antibody, which is determinative of the presenceof the target in the presence of a heterogeneous population of moleculesincluding biological molecules. For example, an antibody thatspecifically binds to a target (which can be an epitope) is an antibodythat binds this target with greater affinity, avidity, more readily,and/or with greater duration than it binds to other targets. In oneembodiment, the extent of binding of an antibody to an unrelated targetis less than about 10% of the binding of the antibody to the target asmeasured, e.g., by a radioimmunoassay (RIA). In certain embodiments, anantibody that specifically binds to a target has a dissociation constant(Kd) of <I μM, <100 nM, <10 nM, <1 nM, or <0.1 nM. In certainembodiments, an antibody specifically binds to an epitope on a proteinthat is conserved among the protein from different species. In anotherembodiment, specific binding can include, but does not require exclusivebinding.

The abbreviations “vc” and “val-cit” refer to the dipeptidevaline-citrulline.

The abbreviation “PAB” refers to the self-immolative spacer:

The abbreviation “MC” refers to the stretcher maleimidocaproyl:

The term “cAC10-MC-vc-PAB-MMAE” refers to a chimeric AC10 antibodyconjugated to the drug MMAE through a MC-vc-PAB linker.

An “anti-CD30 vc-PAB-MMAE antibody-drug conjugate” refers to ananti-CD30 antibody conjugated to the drug MMAE via a linker comprisingthe dipeptide valine citrulline and the self-immolative spacer PAB asshown in Formula (I) of U.S. Pat. No. 9,211,319.

A “cancer” refers a broad group of various diseases characterized by theuncontrolled growth of abnormal cells in the body. A “cancer” or “cancertissue” can include a tumor. Unregulated cell division and growthresults in the formation of malignant tumors that invade neighboringtissues and can also metastasize to distant parts of the body throughthe lymphatic system or bloodstream. Following metastasis, the distaltumors can be said to be “derived from” the pre-metastasis tumor. Forexample, a “tumor derived from” a non-Hodgkin lymphoma refers to a tumorthat is the result of a metastasized non-Hodgkin lymphoma. Because thedistal tumor is derived from the pre-metastasis tumor, the “derivedfrom” tumor can also comprise the pre-metastasis tumor, e.g., a tumorderived from a non-Hodgkin lymphoma can comprise a non-Hodgkin lymphoma.

“CD30” or “TNFRSF8” refers to a receptor that is a member of the tumornecrosis factor receptor superfamily. CD30 is a transmembraneglycoprotein expressed on activated CD4⁺ and CD8⁺ T cells and B cells,and virally-infected lymphocytes. CD30 interacts with TRAF2 and TRAF3 tomediate signal transduction that leads to activation of NF-κB. CD30 actsas a positive regulator of apoptosis, and it has been shown to limit theproliferative potential of auto-reactive CD8 effector T cells. CD30 isalso expressed by various forms of lymphoma, including Hodgkin lymphoma(CD30 is expressed by Reed-Sternberg cells) and non-Hodgkin lymphoma(e.g., diffuse large B-cell lymphoma (DLBCL), peripheral T-cell lymphoma(PTCL), and cutaneous T-cell lymphoma (CTCL).

The terms “Treg” or “regulatory T cell” refer to CD4⁺ T cells thatsuppresses CD4 CD25⁺ and CD8⁺ T cell proliferation and/or effectorfunction, or that otherwise down-modulate an immune response. Notably,Treg may down-regulate immune responses mediated by Natural Killercells, Natural Killer T cells as well as other immune cells.

The terms “regulatory T cell function” or “a function of Treg” are usedinterchangeably to refer to any biological function of a Treg thatresults in a reduction in CD4 CD25⁺ or CD8⁺ T cell proliferation or areduction in an effector T cell-mediated immune response. Treg functioncan be measured via techniques established in the art. Non-limitingexamples of useful in vitro assays for measuring Treg function includeTranswell suppression assays as well as in vitro assays in which thetarget conventional T cells (Tconv) and Tregs purified from humanperipheral blood or umbilical cord blood (or murine spleens or lymphnodes) are optionally activated by anti-CD3⁺ anti-CD28 coated beads (orantigen-presenting cells (APCs) such as, e.g., irradiated splenocytes orpurified dendritic cells (DCs) or irradiated PBMCs) followed by in vitrodetection of conventional T cell proliferation (e.g., by measuringincorporation of radioactive nucleotides (such as, e.g., [H]-thynidine)or fluorescent nucleotides, or by Cayman Chemical MTT Cell ProliferationAssay Kit, or by monitoring the dilution of a green fluorochrome esterCFSE or Seminaphtharhodafluor (SNARF-1) dye by flow cytometry). Othercommon assays measure T cell cytokine responses. Useful in vivo assaysof Treg function include assays in animal models of diseases in whichTregs play an important role, including, e.g., (1) homeostasis model(using naïve homeostatically expanding CD4⁺ T cells as target cells thatare primarily suppressed by Tregs), (2) inflammatory bowel disease (IBD)recovery model (using Th1 T cells (Th17) as target cells that areprimarily suppressed by Tregs), (3) experimental autoimmuneencephalomyelitis (EAE) model (using Th17 and Th1 T cells as targetcells that are primarily suppressed by Tregs), (4) B16 melanoma model(suppression of antitumor immunity) (using CD8⁺ T cells as target cellsthat are primarily suppressed by Tregs), (5) suppression of coloninflammation in adoptive transfer colitis where naïve CD4⁺ CD45RB^(M)Tconv cells are transferred into RagV mice, and (6) Foxp3 rescue model(using lymphocytes as target cells that are primarily suppressed byTregs). According to one protocol, all of the models require mice fordonor T cell populations as well as Ragl^(−/−) or Foxp3 mice forrecipients. For more details on various useful assays see, e.g.,Collison and Vignali, In Vitro Treg Suppression Assays, Chapter 2 inRegulatory T Cells: Methods and Protocols, Methods in Molecular Biology,Kassiotis and Liston eds., Springer, 2011, 707:21-37; Workman et al, InVivo Treg Suppression Assays, Chapter 9 in Regulatory T Cells: Methodsand Protocols, Methods in Molecular Biology, Kassiotis and Liston eds.,Springer, 2011, 119-156; Takahashi et al, Int. Immunol, 1998, 10:1969-1980; Thornton et al, J. Exp. Med., 1998, 188:287-296; Collison etal, J. Immunol, 2009, 182:6121-6128; Thornton and Shevach. J. Exp. Med.,1998, 188:287-296; Assenan et al, J. Exp. Med., 1999, 190:995-1004;Dieckmann et al, J. Exp. Med., 2001, 193: 1303-1310; Belkaid, NatureReviews, 2007, 7:875-888; Tang and Bluestone, Nature Immunology, 2008,9:239-244; Bettini and Vignali, Curr. Opin. Immunol, 2009, 21:612-618;Dannull et al, J Clin Invest. 2005, 115(12):3623-33; Tsaknaridis, et al,J Neurosci Res., 2003, 74:296-308.

The term “immunotherapy” refers to the treatment of a subject afflictedwith, at risk of contracting, or suffering a recurrence of a disease bya method comprising inducing, enhancing, suppressing, or otherwisemodifying an immune response.

“Treatment” or “therapy” of a subject refers to any type of interventionor process performed on, or the administration of an active agent to,the subject with the objective of reversing, alleviating, ameliorating,inhibiting, slowing down, or preventing the onset, progression,development, severity, or recurrence of a symptom, complication,condition, or biochemical indicia associated with a disease.

A “subject” includes any human or non-human animal. The term “nonhumananimal” includes, but is not limited to, vertebrates such as nonhumanprimates, sheep, dogs, and rodents such as mice, rats, and guinea pigs.In some embodiments, the subject is a human. The terms “subject” and“patient” and “individual” are used interchangeably herein.

A “therapeutically effective amount” or “therapeutically effectivedosage” of a drug or therapeutic agent is any amount of the drug that,when used alone or in combination with another therapeutic agent,protects a subject against the onset of a disease or promotes diseaseregression evidenced by a decrease in severity of disease symptoms, anincrease in frequency and duration of disease symptom-free periods, or aprevention of impairment or disability due to the disease affliction.The ability of a therapeutic agent to promote disease regression can beevaluated using a variety of methods known to the skilled practitioner,such as in human subjects during clinical trials, in animal modelsystems predictive of efficacy in humans, or by assaying the activity ofthe agent in in vitro assays.

As used herein, “subtherapeutic dose” means a dose of a therapeuticcompound (e.g., an antibody) that is lower than the usual or typicaldose of the therapeutic compound when administered alone for thetreatment of a hyperproliferative disease (e.g., cancer).

By way of example, an “anti-cancer agent” promotes cancer regression ina subject. In some embodiments, a therapeutically effective amount ofthe drug promotes cancer regression to the point of eliminating thecancer. “Promoting cancer regression” means that administering aneffective amount of the drug, alone or in combination with ananti-cancer agent, results in a reduction in tumor growth or size,necrosis of the tumor, a decrease in severity of at least one diseasesymptom, an increase in frequency and duration of disease symptom-freeperiods, or a prevention of impairment or disability due to the diseaseaffliction. In addition, the terms “effective” and “effectiveness” withregard to a treatment includes both pharmacological effectiveness andphysiological safety. Pharmacological effectiveness refers to theability of the drug to promote cancer regression in the patient.Physiological safety refers to the level of toxicity or other adversephysiological effects at the cellular, organ and/or organism level(adverse effects) resulting from administration of the drug.

By way of example for the treatment of tumors, a therapeuticallyeffective amount of an anti-cancer agent inhibits cell growth or tumorgrowth by at least about 10%, by at least about 20%, by at least about30%, by at least about 40%, by at least about 50%, by at least about60%, by at least about 70%, or by at least about 80%, by at least about90%, at least about 95%, or at least about 100% relative to untreatedsubjects.

In other embodiments of the disclosure, tumor regression can be observedand continue for a period of at least about 20 days, at least about 30days, at least about 40 days, at least about 50 days, or at least about60 days. Notwithstanding these ultimate measurements of therapeuticeffectiveness, evaluation of immunotherapeutic drugs must also makeallowance for “immune-related response patterns”.

An “immune-related response pattern” refers to a clinical responsepattern often observed in cancer patients treated with immunotherapeuticagents that produce antitumor effects by inducing cancer-specific immuneresponses or by modifying native immune processes. This response patternis characterized by a beneficial therapeutic effect that follows aninitial increase in tumor burden or the appearance of new lesions, whichin the evaluation of traditional chemotherapeutic agents would beclassified as disease progression and would be synonymous with drugfailure. Accordingly, proper evaluation of immunotherapeutic agents canrequire long-term monitoring of the effects of these agents on thetarget disease.

“Sustained response” refers to the sustained effect on reducing tumorgrowth after cessation of a treatment. For example, the tumor size mayremain to be the same or smaller as compared to the size at thebeginning of the administration phase. In some embodiments, thesustained response has a duration at least the same as the treatmentduration, at least 1.5×, 2. OX, 2.5×, or 3. OX length of the treatmentduration.

As used herein, “complete response” or “CR” refers to disappearance ofall target lesions; “partial response” or “PR” refers to at least a 30%decrease in the sum of the longest diameters (SLD) of target lesions,taking as reference the baseline SLD; and “stable disease” or “SD”refers to neither sufficient shrinkage of target lesions to qualify forPR, nor sufficient increase to qualify for PD, taking as reference thesmallest SLD since the treatment started.

As used herein, “progression free survival” (PFS) refers to the lengthof time during and after treatment during which the disease beingtreated (e.g., cancer) does not get worse. Progression-free survival mayinclude the amount of time patients have experienced a complete responseor a partial response, as well as the amount of time patients haveexperienced stable disease.

As used herein, “overall response rate” (ORR) refers to the sum ofcomplete response (CR) rate and partial response (PR) rate.

As used herein, “overall survival” refers to the percentage ofindividuals in a group who are likely to be alive after a particularduration of time.

A therapeutically effective amount of a drug includes a“prophylactically effective amount,” which is any amount of the drugthat, when administered alone or in combination with an anti-canceragent to a subject at risk of developing a cancer (e.g., a subjecthaving a pre-malignant condition) or of suffering a recurrence ofcancer, inhibits the development or recurrence of the cancer. In someembodiments, the prophylactically effective amount prevents thedevelopment or recurrence of the cancer entirely. “Inhibiting” thedevelopment or recurrence of a cancer means either lessening thelikelihood of the cancer's development or recurrence, or preventing thedevelopment or recurrence of the cancer entirely.

The term “weight-based dose”, as referred to herein, means that a doseadministered to a patient is calculated based on the weight of thepatient. For example, when a patient with 60 kg body weight requires 3mg/kg of an anti-CD30 antibody, one can calculate and use theappropriate amount of the anti-CD30 antibody (i.e., 180 mg) foradministration.

The use of the term “flat dose” with regard to the methods and dosagesof the disclosure means a dose that is administered to a patient withoutregard for the weight or body surface area (BSA) of the patient. Theflat dose is therefore not provided as a mg/kg dose, but rather as anabsolute amount of the agent (e.g., the anti-CD30 antibody). Forexample, a 60 kg person and a 100 kg person would receive the same doseof an antibody (e.g., 240 mg of an anti-CD30 antibody).

The phrase “pharmaceutically acceptable” indicates that the substance orcomposition must be compatible chemically and/or toxicologically, withthe other ingredients comprising a formulation, and/or the mammal beingtreated therewith.

The phrase “pharmaceutically acceptable salt” as used herein, refers topharmaceutically acceptable organic or inorganic salts of a compound ofthe invention. Exemplary salts include, but are not limited, to sulfate,citrate, acetate, oxalate, chloride, bromide, iodide, nitrate,bisulfate, phosphate, acid phosphate, isonicotinate, lactate,salicylate, acid citrate, tartrate, oleate, tannate, pantothenate,bitartrate, ascorbate, succinate, maleate, gentisinate, fumarate,gluconate, glucuronate, saccharate, formate, benzoate, glutamate,methanesulfonate “mesylate”, ethanesulfonate, benzenesulfonate,/?-toluenesulfonate, pamoate (i.e.,I,Γ-methylene-bis-(2-hydroxy-3-naphthoate)) salts, alkali metal (e.g.,sodiun and potassium) salts, alkaline earth metal (e.g., magnesium)salts, and ammonium salts. A pharmaceutically acceptable salt mayinvolve the inclusion of another molecule such as an acetate ion, asuccinate ion or other counter ion. The counter ion may be any organicor inorganic moiety that stabilizes the charge on the parent compound.Furthermore, a pharmaceutically acceptable salt may have more than onecharged atom in its structure. Instances where multiple charged atomsare part of the pharmaceutically acceptable salt can have multiplecounter ions. Hence, a pharmaceutically acceptable salt can have one ormore charged atoms and/or one or more counter ion.

The use of the alternative (e.g., “or”) should be understood to meaneither one, both, or any combination thereof of the alternatives. Asused herein, the indefinite articles “a” or “an” should be understood torefer to “one or more” of any recited or enumerated component.

The terms “about” or “comprising essentially of” refer to a value orcomposition that is within an acceptable error range for the particularvalue or composition as determined by one of ordinary skill in the art,which will depend in part on how the value or composition is measured ordetermined, i.e., the limitations of the measurement system. Forexample, “about” or “comprising essentially of” can mean within 1 ormore than 1 standard deviation per the practice in the art.Alternatively, “about” or “comprising essentially of” can mean a rangeof up to 20%. Furthermore, particularly with respect to biologicalsystems or processes, the terms can mean up to an order of magnitude orup to 5-fold of a value. When particular values or compositions areprovided in the application and claims, unless otherwise stated, themeaning of “about” or “comprising essentially of” should be assumed tobe within an acceptable error range for that particular value orcomposition.

The terms “once about every week,” “once about every two weeks,” or anyother similar dosing interval terms as used herein mean approximatenumbers. “Once about every week” can include every seven days±one day,i.e., every six days to every eight days. “Once about every two weeks”can include every fourteen days±three days, i.e., every eleven days toevery seventeen days. Similar approximations apply, for example, to onceabout every three weeks, once about every four weeks, once about everyfive weeks, once about every six weeks, and once about every twelveweeks. In some embodiments, a dosing interval of once about every sixweeks or once about every twelve weeks means that the first dose can beadministered any day in the first week, and then the next dose can beadministered any day in the sixth or twelfth week, respectively. Inother embodiments, a dosing interval of once about every six weeks oronce about every twelve weeks means that the first dose is administeredon a particular day of the first week (e.g., Monday) and then the nextdose is administered on the same day of the sixth or twelfth weeks(i.e., Monday), respectively.

As described herein, any concentration range, percentage range, ratiorange, or integer range is to be understood to include the value of anyinteger within the recited range and, when appropriate, fractionsthereof (such as one tenth and one hundredth of an integer), unlessotherwise indicated.

Various aspects of the disclosure are described in further detail in thefollowing subsections.

II. Methods of the Invention

In one aspect, the methods disclosed herein are used in place ofstandard of care therapies. The anti-CD30 antibody-drug conjugatesdescribed herein are used to decrease the activity of CD30⁺ T regulatorycells and/or increase the ratio of CD8⁺ T cells to CD30⁺ T regulatorycells in subjects having cancer, which can result in improved treatmentcompared to standard of care therapies. In certain embodiments, astandard of care therapy is used in combination with any methoddisclosed herein. Standard-of-care therapies for different types ofcancer are well known by persons of skill in the art. For example, theNational Comprehensive Cancer Network (NCCN), an alliance of 21 majorcancer centers in the USA, publishes the NCCN Clinical PracticeGuidelines in Oncology (NCCN GUIDELINES®) that provide detailedup-to-date information on the standard-of-care treatments for a widevariety of cancers (see NCCN GUIDELINES®, 2014, available at:www.nccn.org/professionals/physician_gls/f_guidelines.asp, last accessedMay 14, 2014).

In some embodiments, the therapy of the present disclosure can be usedto treat a lymphoma (e.g., a tumor derived from a lymphoma). Lymphoma isa form of cancer that affects the immune system. The majority oflymphomas fall within two categories: Hodgkin lymphoma (HL) andnon-Hodgkin lymphoma (NHL). NHL is the most common form of lymphoma,accounting for about 90% of all cases of lymphoma, whereas HL accountsfor only about 10% of all cases of lymphoma. Accordingly, in someembodiments of the methods provided herein, the lymphoma is an HL. Inother embodiments of the methods provided herein, the lymphoma is anNHL.

NHL will account for an estimated 72,000 new cases (4.3% of all newcancer cases) and 20,000 deaths (3.4% of all cancer-related deaths) inthe U.S. in 2017. See Howlader N et al., SEER Cancer Statistics Review,1975-2014, based on November 2016 SEER data submission. Diffuse largeB-cell lymphoma (DLBCL), the most common NHL subtype, has an incidencerate of 7.14 per 100,000 persons per year (P-Y), including up to 10%primary mediastinal B-cell lymphoma (PMBL). See Dunleavy K et al., Blood2015; 125:33-39. Incidence rates of peripheral T-cell lymphoma (PTCL)and mycosis fungoides/Sézary syndrome (MF/SS) are 0.60 and 0.52 per100,000 P-Y. See Morton L M et al., Blood 2006; 107:265-276. Within thetwo main categories of lymphoma, HL and NHL, there are several specificsubgroups of lymphomas. Hodgkin lymphomas can include, but are notlimited to, classical HL (cHL; e.g., nodular sclerosing HL, mixedcellularity HL, lymphocyte rich HL, and lymphocyte depleted HL) andnodular lymphocyte predominant type HL. Non-Hodgkin Lymphomas caninclude, but are not limited to, B-cell lymphomas (e.g., diffuse largeB-cell lymphoma (DLBCL), follicular lymphoma (FL), Burkitt lymphoma,immunoblastic large cell lymphoma, precursor B-lymphoblastic lymphoma,and mantle cell lymphoma) and T cell lymphomas (e.g., a cutaneous T-celllymphoma (CTCL), a peripheral T-cell lymphoma (PTCL), a mycosisfungoides, an anaplastic large cell lymphoma, and a precursorT-lymphoblastic lymphoma).

Treatment guidelines for relapsed/refractory (R/R) NHL recommendmulti-agent chemotherapy (combined with targeted therapy for B-celllymphomas), brentuximab vedotin (BV), autologous or allogeneichematopoietic stem cell transplantation (HSCT), and/or radiotherapy,with addition of topical therapies for MF/SS. National ComprehensiveCancer Network, Non-Hodgkin Lymphoma (version 3.2016). 5-year relativesurvival rates are 48%, 44%, and 86% in DLBCL, PTCL, and MF/SS,respectively. SeeHan X et al., Cancer Causes Control 2008; 19:841-858.

A. Anti-CD30 Antibody-Drug Conjugates

In one aspect, the therapy of the present disclosure utilizes ananti-CD30 antibody or an antigen-binding fragment thereof. CD30receptors are members of the tumor necrosis factor receptor superfamilyinvolved in limiting the proliferative potential of autoreactive CD8effector T cells. Antibodies targeting CD30 can potentially be eitheragonists or antagonists of these CD30 mediated activities.

Murine anti-CD30 mAbs known in the art have been generated byimmunization of mice with Hodgkin's disease (HD) cell lines or purifiedCD30 antigen. AC10, originally termed C10 (Bowen et al., 1993, J.Immunol. 151:5896 5906), is distinct in that this anti-CD30 mAb that wasprepared against a hum an NK-like cell line, YT (Bowen et al., 1993, J.Immunol. 151:5896 5906). Initially, the signaling activity of this mAbwas evidenced by the down regulation of the cell surface expression ofCD28 and CD45 molecules, the up regulation of cell surface CD25expression and the induction of homotypic adhesion following binding ofC10 to YT cells. Sequences of the AC10 antibody are set out in SEQ IDNO: 1-16 and Table A below. See also U.S. Pat. No. 7,090,843,incorporated herein by reference.

Generally, antibodies of the disclosure immunospecifically bind CD30 andexert cytostatic and cytotoxic effects on malignant cells in Hodgkin'sdisease. Antibodies of the disclosure are preferably monoclonal, and maybe multispecific, human, humanized or chimeric antibodies, single chainantibodies, Fab fragments, F(ab′) fragments, fragments produced by a Fabexpression library, and CD30 binding fragments of any of the above. Theimmunoglobulin molecules of the disclosure can be of any type (e.g.,IgG, IgE, IgM, IgD, IgA and IgY), class (e.g., IgG1, IgG2, IgG3, IgG4,IgA1 and IgA2) or subclass of immunoglobulin molecule.

In certain embodiments of the disclosure, the antibodies are humanantigen-binding antibody fragments of the present disclosure andinclude, but are not limited to, Fab, Fab′ and F(ab′)₂, Fd, single-chainFvs (scFv), single-chain antibodies, disulfide-linked Fvs (sdFv) andfragments comprising either a V_(L) or V_(H) domain. Antigen-bindingantibody fragments, including single-chain antibodies, may comprise thevariable region(s) alone or in combination with the entirety or aportion of the following: hinge region, CH1, CH2, CH3 and CL domains.Also included in the disclosure are antigen-binding fragments alsocomprising any combination of variable region(s) with a hinge region,CH1, CH2, CH3 and CL domains. Preferably, the antibodies are human,murine (e.g., mouse and rat), donkey, sheep, rabbit, goat, guinea pig,camelid, horse, or chicken.

The antibodies of the present disclosure may be monospecific,bispecific, trispecific or of greater multi specificity. Multispecificantibodies may be specific for different epitopes of CD30 or may bespecific for both CD30 as well as for a heterologous protein. See, e.g.,PCT publications WO 93/17715; WO 92/08802; WO 91/00360; WO 92/05793;Tutt, et al., 1991, J. Immunol. 147:60 69; U.S. Pat. Nos. 4,474,893;4,714,681; 4,925,648; 5,573,920; 5,601,819; Kostelny et al., 1992, J.Immunol. 148:1547 1553.

Antibodies of the present disclosure may be described or specified interms of the particular CDRs they comprise. In certain embodimentsantibodies of the disclosure comprise one or more CDRs of AC10. Thedisclosure encompasses an antibody or derivative thereof comprising aheavy or light chain variable domain, said variable domain comprising(a) a set of three CDRs, in which said set of CDRs are from monoclonalantibody AC10, and (b) a set of four framework regions, in which saidset of framework regions differs from the set of framework regions inmonoclonal antibody AC 10, and in which said antibody or derivativethereof immunospecifically binds CD30.

In one aspect, the anti-CD30 antibody is AC10. In some embodiments, theanti-CD30 antibody is cAC10. cAC10 is a chimeric IgG1 monoclonalantibody that specifically binds CD30. cAC10 induces growth arrest ofCD30⁺ cell lines in vitro and has pronounced antitumor activity insevere combined immunodeficiency (SCID) mouse xenograft models ofHodgkin disease. See Francisco et al., Blood 102(4):1458-64 (2003). AC10antibody and cAC10 antibody are described in U.S. Pat. Nos. 9,211,319,7,090,843.

In one aspect, anti-CD30 antibodies that compete with AC10 antibodyand/or cAC10 antibody binding to CD30 are provided. Anti-CD30 antibodiesthat bind to the same epitope as AC10 antibody and cAC10 antibody arealso provided.

In one aspect, provided herein is an anti-CD30 antibody comprising 1, 2,3, 4, 5, or 6 of the CDR sequences of the AC10 antibody. In one aspect,provided herein is an anti-CD30 antibody comprising 1, 2, 3, 4, 5, or 6of the CDR sequences of the cAC10 antibody. In some embodiments, the CDRis a Kabat CDR or a Chothia CDR.

In one aspect, provided herein is an anti-CD30 antibody comprising aheavy chain variable region and a light chain variable region, whereinthe heavy chain variable region comprises (i) CDR-H1 comprising theamino acid sequence of SEQ ID NO:1, (ii) CDR-H2 comprising the aminoacid sequence of SEQ ID NO:2, and (iii) CDR-H3 comprising the amino acidsequence of SEQ ID NO:3; and/or wherein the light chain variable regioncomprises (i) CDR-L1 comprising the amino acid sequence of SEQ ID NO:4,(ii) CDR-L2 comprising the amino acid sequence of SEQ ID NO:5, and (iii)CDR-L3 comprising the amino acid sequence of SEQ ID NO:6.

An anti-CD30 antibody described herein may comprise any suitableframework variable domain sequence, provided that the antibody retainsthe ability to bind CD30 (e.g., human CD30). As used herein, heavy chainframework regions are designated “HC-FR1-FR4,” and light chain frameworkregions are designated “LC-F1FR4.” In some embodiments, the anti-CD30antibody comprises a heavy chain variable domain framework sequence ofSEQ ID NO:9, 10, 11, and 12 (HC-FR1, HC-FR2, HC-FR3, and HC-FR4,respectively). In some embodiments, the anti-CD30 antibody comprises alight chain variable domain framework sequence of SEQ ID NO:13, 14, 15,and 16 (LC-FR1, LC-FR2, LC-FR3, and LC-FR4, respectively).

In one embodiment, an anti-CD30 antibody comprises a heavy chainvariable domain comprising a framework sequence and hypervariableregions, wherein the framework sequence comprises the HC-FR1-HC-FR4amino acid sequences of SEQ ID NO:9 (HC-FR1), SEQ ID NO:10 (HC-FR2), SEQID NO:11 (HC-FR3), and SEQ ID NO:12 (HC-FR4), respectively; the CDR-H1comprises the amino acid sequence of SEQ ID NO:1; the CDR-H2 comprisesthe amino acid sequence of SEQ ID NO:2; and the CDR-H3 comprises theamino acid sequence of SEQ ID NO:3.

In one embodiment, an anti-CD30 antibody comprises a light chainvariable domain comprising a framework sequence and hypervariableregions, wherein the framework sequence comprises the LC-FR1-LC-FR4amino acid sequences of SEQ ID NO:13 (LC-FR1), SEQ ID NO:14 (LC-FR2),SEQ ID NO:15 (LC-FR3), and SEQ ID NO:16 (LC-FR4), respectively; theCDR-L1 comprises the amino acid sequence of SEQ ID NO:4; the CDR-L2comprises the amino acid sequence of SEQ ID NO:5; and the CDR-L3comprises the amino acid sequence of SEQ ID NO:6.

In some embodiments of the anti-CD30 antibodies described herein, theheavy chain variable domain comprises the amino acid sequence ofQIQLQQSGPEVVKPGASVKISCKASGYTFTDYYITWVKQKPGQGLEWIGWIYPGSGNTKYNEKFKGKATLTVDTSSSTAFMQLSSLTSEDTAVYFCANYGNYWFAYWGQGTQ VTVSA (SEQ IDNO:7) and the light chain variable domain comprises the amino acidsequence of DIVLTQSPASLAVSLGQRATISCKASQSVDFDGDSYMNWYQQKPGQPPKVLIYAASNLESGIPARFSGSGSGTDFTLNIHPVEEEDAATYYCQQSNEDPWTFGGGTKLEIK (SEQ ID NO:8).

In some embodiments of the anti-CD30 antibodies described herein, theheavy chain CDR sequences comprise the following:

a) CDR-H1 (DYYIT (SEQ ID NO: 1)); b) CDR-H2(WIYPGSGNTKYNEKFKG (SEQ ID NO: 2)); and c) CDR-H3(YGNYWFAY (SEQ ID NO: 3)).

In some embodiments of the anti-CD30 antibodies described herein, theheavy chain FR sequences comprise the following:

a) HC-FR1 (QIQLQQSGPEVVKPGASVKISCKASGYTFT (SEQ ID NO: 9)); b) HC-FR2(WVKQKPGQGLEWIG (SEQ ID NO: 10)); c) HC-FR3(KATLTVDTSSSTAFMQLSSLTSEDTAVYFCAN (SEQ ID NO: 11)); and d) HC-FR4(WGQGTQVTVSA (SEQ ID NO: 12)).

In some embodiments of the anti-CD30 antibodies described herein, thelight chain CDR sequences comprise the following:

a) CDR-L1 (KASQSVDFDGDSYMN (SEQ ID NO: 4)); b) CDR-L2(AASNLES (SEQ ID NO: 5));  and c) CDR-L3 (QQSNEDPWT (SEQ ID NO: 6)).

In some embodiments of the anti-CD30 antibodies described herein, thelight chain FR sequences comprise the following:

a) LC-FR1 (DIVLTQSPASLAVSLGQRATISC (SEQ ID NO: 13)); b) LC-FR2(WYQQKPGQPPKVLIY (SEQ ID NO: 14)); c) LC-FR3(GIPARFSGSGSGTDFTLNIHPVEEEDAATYYC (SEQ ID NO: 15));  and d) LC-FR4(FGGGTKLEIK (SEQ ID NO: 16)).

In some embodiments, provided herein is an anti-CD30 antibody that bindsto CD30 (e.g., human CD30), wherein the antibody comprises a heavy chainvariable region and a light chain variable region, wherein the antibodycomprises:

-   -   (a) heavy chain variable domain comprising:    -   (1) an HC-FR1 comprising the amino acid sequence of SEQ ID NO:9;    -   (2) an CDR-H1 comprising the amino acid sequence of SEQ ID NO:1;    -   (3) an HC-FR2 comprising the amino acid sequence of SEQ ID        NO:10;    -   (4) an CDR-H2 comprising the amino acid sequence of SEQ ID NO:2;    -   (5) an HC-FR3 comprising the amino acid sequence of SEQ ID NO:        11;    -   (6) an CDR-H3 comprising the amino acid sequence of SEQ ID NO:3;        and    -   (7) an HC-FR4 comprising the amino acid sequence of SEQ ID        NO:12, and/or    -   (b) a light chain variable domain comprising:    -   (1) an LC-FR1 comprising the amino acid sequence of SEQ ID        NO:13;    -   (2) an CDR-L1 comprising the amino acid sequence of SEQ ID NO:4;    -   (3) an LC-FR2 comprising the amino acid sequence of SEQ ID        NO:14;    -   (4) an CDR-L2 comprising the amino acid sequence of SEQ ID NO:5;    -   (5) an LC-FR3 comprising the amino acid sequence of SEQ ID        NO:15;    -   (6) an CDR-L3 comprising the arnino acid sequence of SEQ ID        NO:6; and    -   (7) an LC-FR4 comprising the amino acid sequence of SEQ ID        NO:16.

In one aspect, provided herein is an anti-CD30 antibody comprising aheavy chain variable domain comprising the amino acid sequence of SEQ IDNO:7 and/or comprising a light chain variable domain comprising theamino acid sequence of SEQ ID NO:8.

In some embodiments, provided herein is an anti-CD30 antibody comprisinga heavy chain variable domain comprising an amino acid sequence havingat least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, or 99% sequence identity to the amino acid sequence of SEQ IDNO:7. In certain embodiments, a heavy chain variable domain comprisingan amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to theamino acid sequence of SEQ ID NO:7 contains substitutions (e.g.,conservative substitutions), insertions, or deletions relative to thereference sequence and retains the ability to bind to a CD30 (e.g.,human CD30). In certain embodiments, a total of 1 to 10 amino acids havebeen substituted, inserted and/or deleted in SEQ ID NO:7. In certainembodiments, substitutions, insertions, or deletions (e.g., 1, 2, 3, 4,or 5 amino acids) occur in regions outside the CDR s (i.e., in the FRs).In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable domain sequence of SEQ ID NO:7 including post-translationalmodifications of that sequence. In a particular embodiment, the heavychain variable domain comprises one, two or three CDRs selected from:(a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:1, (b) CDR-H2comprising the amino acid sequence of SEQ ID NO:2, and (c) CDR-H3comprising the amino acid sequence of SEQ ID NO:3.

In some embodiments, provided herein is an anti-CD30 antibody comprisinga light chain variable domain comprising an amino acid sequence havingat least 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, or 99% sequence identity to the amino acid sequence of SEQ IDNO:8. In certain embodiments, a light chain variable domain comprisingan amino acid sequence having at least 85%, 86%, 87%, 88%, 89%, 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to theamino acid sequence of SEQ ID NO:8 contains substitutions (e.g.,conservative substitutions), insertions, or deletions relative to thereference sequence and retains the ability to bind to a CD30 (e.g.,human CD30). In certain embodiments, a total of 1 to 10 amino acids havebeen substituted, inserted and/or deleted in SEQ ID NO:8. In certainembodiments, substitutions, insertions, or deletions (e.g., 1, 2, 3, 4,or 5 amino acids) occur in regions outside the CDR s (i.e., in the FRs).In some embodiments, the anti-CD30 antibody comprises a light chainvariable domain sequence of SEQ ID NO:8 including post-translationalmodifications of that sequence. In a particular embodiment, the lightchain variable domain comprises one, two or three CDRs selected from:(a) CDR-H1 comprising the amino acid sequence of SEQ ID NO:4, (b) CDR-H2comprising the amino acid sequence of SEQ ID NO:5, and (c) CDR-H3comprising the amino acid sequence of SEQ ID NO:6.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable domain as in any of the embodiments provided above, and a lightchain variable domain as in any of the embodiments provided above. Inone embodiment, the antibody comprises the heavy chain variable domainsequence of SEQ ID NO:7 and the light chain variable domain sequence ofSEQ ID NO:8, including post-translational modifications of thosesequences.

In some embodiments, the anti-CD30 antibody of the anti-CD30antibody-drug conjugate comprises: i) a heavy chain CDR1 set out in SEQID NO: 1, a heavy chain CDR2 set out in SEQ ID NO: 2, a heavy chain CDR3set out in SEQ ID NO: 3; and ii) a light chain CDR1 set out in SEQ IDNO: 4, a light chain CDR2 set out in SEQ ID NO: 5, and a light chainCDR3 set out in SEQ ID NO: 6.

In some embodiments, the anti-CD30 antibody of the anti-CD30antibody-drug conjugate comprises: i) an amino acid sequence at least85% identical to a heavy chain variable region set out in SEQ ID NO: 7,and ii) an amino acid sequence at least 85% identical to a light chainvariable region set out in SEQ ID NO: 8.

In some embodiments, the anti-CD30 antibody of the anti-CD30antibody-drug conjugate is a monoclonal antibody.

In some embodiments, the anti-CD30 antibody of the anti-CD30antibody-drug conjugate is a chimeric AC10 antibody.

Antibodies of the present invention may also be described or specifiedin terms of their binding affinity to CD30. Preferred binding affinitiesinclude those with a dissociation constant or Kd less than 5×10² M, 10⁻²M, 5×10⁻³ M, 10⁻³ M, 5×10⁻⁴ M, 10⁻⁴ M, 5×10⁻⁵ M, 10⁻⁵ M, 5×10⁻⁶ M, 10⁻⁶M, 5×10⁻⁷ M, 10⁻⁷ M, 5×10⁻⁸ M, 10⁻⁸M, 5×10⁻⁹ M, 10⁻⁹ M, 5×10⁻¹⁰ M, 10⁻¹⁰M, 5×10⁻¹¹ M, 10⁻¹¹ M, 5×10⁻¹² M, 10⁻¹² M, 5×10⁻¹³ M, 10⁻¹³ M, 5×10⁻¹⁴M, 10⁻¹⁴ M, 5×10⁻¹⁵ M, or 10⁻¹⁵ M.

There are five classes of immunoglobulins: IgA, IgD, IgE, IgG and IgM,having heavy chains designated α, δ, ε, γ and μ, respectively. The γ andα classes are further divided into subclasses e.g., humans express thefollowing subclasses: IgG1, IgG2, IgG3, IgG4, IgA1 and IgA2. IgG1antibodies can exist in multiple polymorphic variants termed allotypes(reviewed in Jefferis and Lefranc 2009. mAbs Vol 1 Issue 4 1-7) any ofwhich are suitable for use in some of the embodiments herein. Commonallotypic variants in human populations are those designated by theletters a, f, n, z or combinations thereof. In any of the embodimentsherein, the antibody may comprise a heavy chain Fc region comprising ahuman IgG Fc region. In further embodiments, the human IgG Fc regioncomprises a human IgG1.

In one aspect of the invention, polynucleotides encoding anti-CD30antibodies, such as those anti-CD30 antibodies described herein, areprovided. In certain embodiments, vectors comprising polynucleotidesencoding anti-CD30 antibodies as described herein are provided. Incertain embodiments, host cells comprising such vectors are provided. Inanother aspect of the invention, compositions comprising anti-CD30antibodies described herein or polynucleotides encoding anti-CD30antibodies described herein are provided.

The antibodies also include derivatives that are modified, i.e., by thecovalent attachment of any type of molecule to the antibody such thatcovalent attachment does not prevent the antibody from binding to CD30or from exerting a cytostatic or cytotoxic effect on HD cells. Forexample, but not by way of limitation, the antibody derivatives includeantibodies that have been modified, e.g., by glycosylation, acetylation,PEGylation, phosphylation, amidation, derivatization by knownprotecting/blocking groups, proteolytic cleavage, linkage to a cellularligand or other protein, etc. Any of numerous chemical modifications maybe carried out by known techniques, including, but not limited tospecific chemical cleavage, acetylation, formylation, metabolicsynthesis of tunicamycin, etc. Additionally, the derivative may containone or more non-classical amino acids.

In some embodiments, the anti-CD30 antibody is conjugated to atherapeutic agent (e.g., an anti-CD30 antibody-drug conjugate). In someembodiments, the therapeutic agent comprises an anti-neoplastic agent(e.g., an anti-mitotic agent). In certain embodiments, the therapeuticagent is selected from the group consisting of monomethyl auristatin E(MMAE), monomethyl auristatin F (MMAF), auristatin drug analogues,cantansinoids, maytansinoids (e.g., maytansine; DMs), dolastatins,cryptophycin, duocarmycin, duocarmycin derivatives, esperamicin,calicheamicin, pyrolobenodiazepine (PBD), and any combination thereof.In one particular embodiment, the anti-CD30 antibody is conjugated toMMAE. The antibody can be conjugated to at least one, at least two, atleast three, at least four, at least five, at least six, at least seven,at least eight, at least nine, or at least ten molecules of thetherapeutic agent (e.g., MMAE). In one embodiment, the anti-CD30antibody is conjugated to four molecules of the therapeutic agent, e.g.,four molecules of MMAE. In one particular embodiment, the anti-CD30antibody is conjugated to MMAF. The antibody can be conjugated to atleast one, at least two, at least three, at least four, at least five,at least six, at least seven, at least eight, at least nine, or at leastten molecules of the therapeutic agent (e.g., MMAF). In one embodiment,the anti-CD30 antibody is conjugated to four molecules of thetherapeutic agent, e.g., four molecules of MMAF.

In some embodiments, the anti-CD30 antibody-drug conjugate furthercomprises a linker between the therapeutic agent and the antibody. Insome embodiments, the linker comprises one or more naturally occurringamino acids, one or more non-naturally occurring (e.g., synthetic) aminoacids, a chemical linker, or any combination thereof. In certainembodiments, the linker is a cleavable linker, e.g., a proteasecleavable linker. In certain embodiments, the linker is specificallycleaved upon uptake by a target cell, e.g., upon uptake by a cellexpressing CD30. In certain embodiments, the linker is a cleavablepeptide linker having the formula: “-MC-vc-PAB-” or “-MC-val-cit-PAB-”,wherein “MC” refers to the stretcher maleimidocaproyl having thefollowing structure:

“vc” and “val-cit” refer to the dipeptide valine-citrulline, and PABrefers to a self-immolative spacer having the following structure:

In some embodiments, cleavage of the linker activates a cytotoxicactivity of the therapeutic agent. In certain embodiments, the linker isa non-cleavable linker. In certain embodiments, the non-cleavable linkerhas the formula: “-MC-”, wherein “MC” refers to the stretchermaleimidocaproyl having the following structure:

In some embodiments, the antibody-drug conjugates comprises an anti-CD30antibody, covalently linked to MMAE through a vc-PAB linker. In someembodiments, the antibody-drug conjugate is delivered to the subject asa pharmaceutical composition. In some embodiments, the CD30 antibodydrug conjugates contemplated herein are as described in U.S. Pat. No.9,211,319, herein incorporated by reference.

In one embodiment, the anti-CD30 antibody drug-conjugate comprisesbrentuximab vedotin. In one particular embodiment, the anti-CD30antibody drug-conjugate is brentuximab vedotin. Brentuximab vedotin (BV;also known as “ADCETRIS®”) is a CD30-directed antibody-drug conjugate(ADC) comprising a chimeric anti-CD30 antibody (cAC10), a therapeuticagent (MMAE), and a protease-cleavable linker between the cAC10 and theMMAE, as shown in the following structure:

The drug to antibody ratio or drug loading is represented by “p” in thestructure of brentuximab vedotin and ranges in integer values from 1 to8. The average drug loading of brentuximab vedotin in a pharmaceuticalcomposition is about 4. ADCETRIS® is approved by the FDA for treatmentof patients with Hodgkin lymphoma after failure of autologous stem celltransplant (ASCT) or after failure of at least two prior multi-agentchemotherapy regimens in patients who are not ASCT candidates and forthe treatment of patients with systemic anaplastic large cell lymphomaafter failure of at least one prior multi-agent chemotherapy regimen.

In one embodiment, the anti-CD30 antibody is an anti-CD30 antibody orantigen-binding fragment thereof that binds to the same epitope ascAC10, e.g., the same epitope as brentuximab vedotin. In certainembodiments, the anti-CD30 antibody is an antibody that has the sameCDRs as cAC10, e.g., the same CDRs as brentuximab vedotin. Antibodiesthat bind to the same epitope are expected to have functional propertiesvery similar to those of cAC10 by virtue of their binding to the sameepitope region of CD30. These antibodies can be readily identified basedon their ability to, for example, cross-compete with cAC10 in standardCD30 binding assays such as Biacore analysis, ELISA assays, or flowcytometry.

In certain embodiments, the antibodies that cross-compete for binding tohuman CD30 with, or bind to the same epitope region of human CD30 ascAC10 are monoclonal antibodies. For administration to human subjects,these cross-competing antibodies can be chimeric antibodies, or can behumanized or human antibodies. Such chimeric, humanized, or humanmonoclonal antibodies can be prepared and isolated by methods well knownin the art. Anti-CD30 antibodies usable in the methods of the discloseddisclosure also include antigen-binding portions of the aboveantibodies.

In other embodiments, the anti-CD30 antibody or antigen-binding portionthereof is a chimeric, humanized, or human monoclonal antibody or aportion thereof. In certain embodiments for treating a human subject,the antibody is a humanized antibody. In other embodiments for treatinga human subject, the antibody is a human antibody. Antibodies of anIgG1, IgG2, IgG3, or IgG4 isotype can be used.

B. Methods of Modulating the Immune Response

In one aspect, the present invention provides for a method of decreasingthe activity of CD30⁺ T regulatory (Treg) cells in a subject havingcancer comprising administering to the subject an antibody-drugconjugate, wherein the antibody-drug conjugate comprises an anti-CD30antibody or an antigen-binding portion thereof conjugated to amonomethyl auristatin.

In some embodiments, decreasing the activity of CD30⁺ Treg cellscomprises a decrease in the number of CD30⁺ Treg cells. In someembodiments, the number of CD30⁺ Treg cells is decreased relative to thenumber of one or more other types of CD4⁺ T cells. In some embodiments,the one or more other types of CD4⁺ T cells comprise Th1 cells, Th2cells or Th17 cells. In some embodiments, the one or more other types ofCD4⁺ T cells comprise Th1 CD30⁺ cells, Th2 CD30⁺ cells or Th17 CD30⁺cells. In some embodiments, the number of CD30⁺ Treg cells is decreasedrelative to the number of CD30⁺ Treg cells in the subject prior toadministration of the antibody-drug conjugate. In some embodiments,number of CD30⁺ Treg cells is decreased relative to the number of CD30⁺Treg cells in a subject who has not been treated with the antibody-drugconjugate.

In some embodiments, decreasing the activity of CD30⁺ Treg cellscomprises a decrease in the function of CD30⁺ Treg cells. In someembodiments, the decrease in the function of CD30⁺ Treg cells isrelative to the function of CD30⁺ Treg cells in a subject prior toadministration of the antibody-drug conjugate. In some embodiments, thedecrease in the function of CD30⁺ Treg cells is relative to the functionof CD30⁺ Treg cells in a subject who has not been treated with theantibody-drug conjugate.

In some embodiments, the CD30⁺ Treg cells are CD30⁺ inducible Tregulatory (iTreg) cells or CD30⁺ peripheral T regulatory (pTreg) cells.

In some embodiments, the monomethyl auristatin is monomethyl auristatinE (MMAE). In some embodiments, the monomethyl auristatin is monomethylauristatin F (MMAF).

In some embodiments, the anti-CD30 antibody is monoclonal anti-CD30antibody AC10. In some embodiments, the anti-CD30 antibody is cAC10. Insome embodiments, the antibody-drug conjugate is brentuximab vedotin.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region and a light chain variable region, wherein the heavychain variable region comprises:

-   -   (i) a CDR-H1 comprising the amino acid sequence of SEQ ID NO: 1;    -   (ii) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:        2; and    -   (iii) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:        3; and        wherein the light chain variable region comprises:    -   (i) a CDR-L1 comprising the amino acid sequence of SEQ ID NO: 4;    -   (ii) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:        5; and    -   (iii) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:        6.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region comprising the amino acid sequence of SEQ ID NO: 7 and alight chain variable region comprising the amino acid sequence of SEQ IDNO: 8.

In some embodiments, the antibody-drug conjugate further comprises alinker between the anti-CD30 antibody or antigen-binding portion thereofand the monomethyl auristatin. In some embodiments, the linker isselected from the group consisting of a cleavable linker and anon-cleavable linker. In some embodiments, the linker is a cleavablepeptide linker. In some embodiments, the cleavable peptide linker has aformula: -MC-vc-PAB-. In some embodiments, the linker is a non-cleavablelinker having a formula: -MC-.

In some embodiments, the subject has been previously treated for thecancer. In some embodiments, the subject did not respond to treatment orrelapsed after first-line treatment. In some embodiments, the subjecthas not previously been treated for the cancer.

In some embodiments, the cancer is a lymphoma. In some embodiments, thelymphoma is a T-cell lymphoma. In some embodiments, the lymphoma is aB-cell lymphoma.

In some embodiments, the lymphoma is a non-Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the non-Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject has not beenpreviously treated for the non-Hodgkin lymphoma. In some embodiments,the non-Hodgkin lymphoma is a mature T-cell lymphoma. In someembodiments, the non-Hodgkin lymphoma is diffuse large B-cell lymphoma(DLBCL), peripheral T-cell lymphoma (PTCL), anaplastic large celllymphoma (ALCL) or cutaneous T-cell lymphoma (CTCL). In someembodiments, the non-Hodgkin lymphoma is cutaneous T-cell lymphoma(CTCL). In some embodiments, the cutaneous T-cell lymphoma (CTCL) is amycosis fungoides (MF). In some embodiments, the mycosis fungoides is aCD30-positive mycosis fungoides (MF). In some embodiments, the cutaneousT-cell lymphoma (CTCL) is a primary cutaneous anaplastic large celllymphoma (pcALCL). In some embodiments, the subject has received priorsystemic treatment. In some embodiments, the non-Hodgkin lymphoma isanaplastic large cell lymphoma (ALCL). In some embodiments, theanaplastic large cell lymphoma (ALCL) is a systemic anaplastic largecell lymphoma (sALCL).

In some embodiments, the lymphoma is a Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject relapsed afterautologous stem cell transplant. In some embodiments, the subjectrelapsed after first-line treatment and the subject is ineligible forautologous stem cell transplant. In some embodiments, the subject hasnot been previously treated for the Hodgkin lymphoma. In someembodiments, the Hodgkin lymphoma is classical Hodgkin lymphoma (cHL).In some embodiments, the classical Hodgkin lymphoma (cHL) is advancedcHL. In some embodiments, the subject has been previously treated forcHL. In some embodiments, the subject has not been previously treatedfor cHL.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents capable of modulating the immuneresponse. In some embodiments, the one or more additional therapeuticagents is not an antibody or antigen-binding fragment thereof. In someembodiments, the one or more additional therapeutic agents is anantibody or antigen-binding fragment thereof.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents. In some embodiments, the one or moreadditional therapeutic agents is a chemotherapy regimen consistingessentially of doxorubicin, vinblastine, and dacarbazine (AVD). In someembodiments, the one or more additional therapeutic agents is achemotherapy regimen consisting essentially of Cyclophosphamide,Doxorubicin, and Prednisone (CHP). In some embodiments, the one or moreadditional therapeutic agents is an alkylating agent, an anthracycline,an antibiotic, an antifolate, an antimetabolite, an antitubulin agent,an auristatin, a chemotherapy sensitizer, a DNA minor groove binder, aDNA replication inhibitor, a duocarmycin, an etoposide, a fluorinatedpyrimidine, a lexitropsin, a nitrosourea, a platinol, a purineantimetabolite, a puromycin, a radiation sensitizer, a steroid, ataxane, a topoisomerase inhibitor, and/or a vinca alkaloid. In someembodiments, the one or more additional therapeutic agents is selectedfrom the group consisting of adriamycin, an androgen, anthramycin (AMC),asparaginase, 5-azacytidine, azathioprine, bleomycin, busulfan,buthionine sulfoximine, camptothecin, carboplatin, carmustine (BSNU),CC-1065, chlorambucil, cisplatin, colchicine, cyclophosphamide,cytarabine, cytidine arabinoside, cytochalasin B, dacarbazine,dactinomycin (formerly actinomycin), daunorubicin, decarbazine,docetaxel, doxorubicin, an estrogen, 5-fluordeoxyuridine,5-fluorouracil, gramicidin D, hydroxydaunorubicin, hydroxyurea,idarubicin, ifosfamide, irinotecan, lomustine (CCNU), mechlorethamine,melphalan, 6-mercaptopurine, methotrexate, mithramycin, mitomycin C,mitoxantrone, nitroimidazole, paclitaxel, plicamycin, prednisone,prednisolone, procarbizine, streptozotocin, tenoposide, 6-thioguanine,thioTEPA, topotecan, vinblastine, vincristine, vinorelbine, VP-16 andVM-26. In some embodiments, the one or more additional therapeuticagents is an antibody or antigen-binding fragment thereof.

In some embodiments, the subject has cHL that has not been previouslytreated and the one or more additional therapeutic agents areadriamycin, dacarabazine and vinblastine. In some embodiments, the cHLis advanced cHL.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and the one or more additional therapeuticagents are cyclophosphamide, hydroxydaunorubicin and prednisone.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and the one or more additional therapeuticagents are cyclophosphamide, hydroxydaunorubicin and prednisolone.

In some embodiments, the method further comprises treating the subjectwith irradiation.

In another aspect, the present invention provides for a method ofincreasing the ratio of CD8⁺ T cells to CD30⁺ T regulatory (Treg) cellsin a subject having cancer comprising administering to the subject anantibody-drug conjugate, wherein the antibody-drug conjugate comprisesan anti-CD30 antibody or an antigen-binding portion thereof conjugatedto a monomethyl auristatin. In some embodiments, the ratio of CD8⁺ Tcells to CD30⁺ Treg cells is increased relative to the ratio of CD8⁺ Tcells to CD30⁺ Treg cells in the subject prior to the administration ofthe antibody-drug conjugate.

In some embodiments, the CD30⁺ Treg cells are CD30⁺ inducible Tregulatory (iTreg) cells or CD30⁺ peripheral T regulatory (pTreg) cells.

In some embodiments, the monomethyl auristatin is monomethyl auristatinE (MMAE). In some embodiments, the monomethyl auristatin is monomethylauristatin F (MMAF).

In some embodiments, the anti-CD30 antibody is monoclonal anti-CD30antibody AC10. In some embodiments, the anti-CD30 antibody is cAC10. Insome embodiments, the antibody-drug conjugate is brentuximab vedotin.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region and a light chain variable region, wherein the heavychain variable region comprises:

-   -   (i) a CDR-H1 comprising the amino acid sequence of SEQ ID NO: 1;    -   (ii) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:        2; and    -   (iii) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:        3; and        wherein the light chain variable region comprises:    -   (i) a CDR-L1 comprising the amino acid sequence of SEQ ID NO: 4;    -   (ii) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:        5; and    -   (iii) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:        6.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region comprising the amino acid sequence of SEQ ID NO: 7 and alight chain variable region comprising the amino acid sequence of SEQ IDNO: 8.

In some embodiments, the antibody-drug conjugate further comprises alinker between the anti-CD30 antibody or antigen-binding portion thereofand the monomethyl auristatin. In some embodiments, the linker isselected from the group consisting of a cleavable linker and anon-cleavable linker. In some embodiments, the linker is a cleavablepeptide linker. In some embodiments, the cleavable peptide linker has aformula: -MC-vc-PAB-. In some embodiments, the linker is a non-cleavablelinker having a formula: -MC-.

In some embodiments, the subject has been previously treated for thecancer. In some embodiments, the subject did not respond to treatment orrelapsed after first-line treatment. In some embodiments, the subjecthas not previously been treated for the cancer.

In some embodiments, the cancer is a lymphoma. In some embodiments, thelymphoma is a T-cell lymphoma. In some embodiments, the lymphoma is aB-cell lymphoma.

In some embodiments, the lymphoma is a non-Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the non-Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject has not beenpreviously treated for the non-Hodgkin lymphoma. In some embodiments,the non-Hodgkin lymphoma is a mature T-cell lymphoma. In someembodiments, the non-Hodgkin lymphoma is diffuse large B-cell lymphoma(DLBCL), peripheral T-cell lymphoma (PTCL), anaplastic large celllymphoma (ALCL) or cutaneous T-cell lymphoma (CTCL). In someembodiments, the non-Hodgkin lymphoma is cutaneous T-cell lymphoma(CTCL). In some embodiments, the cutaneous T-cell lymphoma (CTCL) is amycosis fungoides (MF). In some embodiments, the mycosis fungoides is aCD30-positive mycosis fungoides (MF). In some embodiments, the cutaneousT-cell lymphoma (CTCL) is a primary cutaneous anaplastic large celllymphoma (pcALCL). In some embodiments, the subject has received priorsystemic treatment. In some embodiments, the non-Hodgkin lymphoma isanaplastic large cell lymphoma (ALCL). In some embodiments, theanaplastic large cell lymphoma (ALCL) is a systemic anaplastic largecell lymphoma (sALCL).

In some embodiments, the lymphoma is a Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject relapsed afterautologous stem cell transplant. In some embodiments, the subjectrelapsed after first-line treatment and the subject is ineligible forautologous stem cell transplant. In some embodiments, the subject hasnot been previously treated for the Hodgkin lymphoma. In someembodiments, the Hodgkin lymphoma is classical Hodgkin lymphoma (cHL).In some embodiments, the classical Hodgkin lymphoma (cHL) is advancedcHL. In some embodiments, the subject has been previously treated forcHL. In some embodiments, the subject has not been previously treatedfor cHL.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents capable of modulating the immuneresponse. In some embodiments, the one or more additional therapeuticagents is not an antibody or antigen-binding fragment thereof. In someembodiments, the one or more additional therapeutic agents is anantibody or antigen-binding fragment thereof.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents. In some embodiments, the one or moreadditional therapeutic agents is a chemotherapy regimen consistingessentially of doxorubicin, vinblastine, and dacarbazine (AVD). In someembodiments, the one or more additional therapeutic agents is achemotherapy regimen consisting essentially of Cyclophosphamide,Doxorubicin, and Prednisone (CHP). In some embodiments, the one or moreadditional therapeutic agents is an alkylating agent, an anthracycline,an antibiotic, an antifolate, an antimetabolite, an antitubulin agent,an auristatin, a chemotherapy sensitizer, a DNA minor groove binder, aDNA replication inhibitor, a duocarmycin, an etoposide, a fluorinatedpyrimidine, a lexitropsin, a nitrosourea, a platinol, a purineantimetabolite, a puromycin, a radiation sensitizer, a steroid, ataxane, a topoisomerase inhibitor, and/or a vinca alkaloid. In someembodiments, the one or more additional therapeutic agents is selectedfrom the group consisting of adriamycin, an androgen, anthramycin (AMC),asparaginase, 5-azacytidine, azathioprine, bleomycin, busulfan,buthionine sulfoximine, camptothecin, carboplatin, carmustine (BSNU),CC-1065, chlorambucil, cisplatin, colchicine, cyclophosphamide,cytarabine, cytidine arabinoside, cytochalasin B, dacarbazine,dactinomycin (formerly actinomycin), daunorubicin, decarbazine,docetaxel, doxorubicin, an estrogen, 5-fluordeoxyuridine,5-fluorouracil, gramicidin D, hydroxydaunorubicin, hydroxyurea,idarubicin, ifosfamide, irinotecan, lomustine (CCNU), mechlorethamine,melphalan, 6-mercaptopurine, methotrexate, mithramycin, mitomycin C,mitoxantrone, nitroimidazole, paclitaxel, plicamycin, prednisone,prednisolone, procarbizine, streptozotocin, tenoposide, 6-thioguanine,thioTEPA, topotecan, vinblastine, vincristine, vinorelbine, VP-16 andVM-26. In some embodiments, the one or more additional therapeuticagents is an antibody or antigen-binding fragment thereof.

In some embodiments, the subject has cHL that has not been previouslytreated and wherein the one or more additional therapeutic agents areadriamycin, dacarabazine and vinblastine. In some embodiments, the cHLis advanced cHL.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and wherein the one or more additionaltherapeutic agents are cyclophosphamide, hydroxydaunorubicin andprednisone.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and wherein the one or more additionaltherapeutic agents are cyclophosphamide, hydroxydaunorubicin andprednisolone.

In some embodiments, the method further comprises treating the subjectwith irradiation.

In another aspect, the present invention provides for a method ofmodulating the immune response in a subject having cancer comprisingadministering to the subject an antibody-drug conjugate, wherein theantibody-drug conjugate comprises an anti-CD30 antibody or anantigen-binding portion thereof conjugated to a monomethyl auristatin,wherein the modulation comprises increasing the ratio of CD8⁺ T cells toCD30⁺ T regulatory (Treg) cells in the subject. In some embodiments, theratio of CD8⁺ T cells to CD30⁺ Treg cells is increased relative to theratio of CD8⁺ T cells to CD30⁺ Treg cells in the subject prior to theadministration of the antibody-drug conjugate. In some embodiments, theratio of CD8⁺ T cells to CD30⁺ Treg cells is increased relative to theratio of CD8⁺ T cells to CD30⁺ Treg cells in a subject who has not beentreated with the antibody-drug conjugate.

In some embodiments, the CD30⁺ Treg cells are CD30⁺ inducible Tregulatory (iTreg) cells or CD30⁺ peripheral T regulatory (pTreg) cells.

In some embodiments, the monomethyl auristatin is monomethyl auristatinE (MMAE). In some embodiments, the monomethyl auristatin is monomethylauristatin F (MMAF).

In some embodiments, the anti-CD30 antibody is monoclonal anti-CD30antibody AC10. In some embodiments, the anti-CD30 antibody is cAC10. Insome embodiments, the antibody-drug conjugate is brentuximab vedotin.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region and a light chain variable region, wherein the heavychain variable region comprises:

-   -   (i) a CDR-H1 comprising the amino acid sequence of SEQ ID NO: 1;    -   (ii) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:        2; and    -   (iii) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:        3; and        wherein the light chain variable region comprises:    -   (i) a CDR-L1 comprising the amino acid sequence of SEQ ID NO: 4;    -   (ii) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:        5; and    -   (iii) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:        6.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region comprising the amino acid sequence of SEQ ID NO: 7 and alight chain variable region comprising the amino acid sequence of SEQ IDNO: 8.

In some embodiments, the antibody-drug conjugate further comprises alinker between the anti-CD30 antibody or antigen-binding portion thereofand the monomethyl auristatin. In some embodiments, the linker isselected from the group consisting of a cleavable linker and anon-cleavable linker. In some embodiments, the linker is a cleavablepeptide linker. In some embodiments, the cleavable peptide linker has aformula: -MC-vc-PAB-. In some embodiments, the linker is a non-cleavablelinker having a formula: -MC-.

In some embodiments, the subject has been previously treated for thecancer. In some embodiments, the subject did not respond to treatment orrelapsed after first-line treatment. In some embodiments, the subjecthas not previously been treated for the cancer.

In some embodiments, the cancer is a lymphoma. In some embodiments, thelymphoma is a T-cell lymphoma. In some embodiments, the lymphoma is aB-cell lymphoma.

In some embodiments, the lymphoma is a non-Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the non-Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject has not beenpreviously treated for the non-Hodgkin lymphoma. In some embodiments,the non-Hodgkin lymphoma is a mature T-cell lymphoma. In someembodiments, the non-Hodgkin lymphoma is diffuse large B-cell lymphoma(DLBCL), peripheral T-cell lymphoma (PTCL), anaplastic large celllymphoma (ALCL) or cutaneous T-cell lymphoma (CTCL). In someembodiments, the non-Hodgkin lymphoma is cutaneous T-cell lymphoma(CTCL). In some embodiments, the cutaneous T-cell lymphoma (CTCL) is amycosis fungoides (MF). In some embodiments, the mycosis fungoides is aCD30-positive mycosis fungoides (MF). In some embodiments, the cutaneousT-cell lymphoma (CTCL) is a primary cutaneous anaplastic large celllymphoma (pcALCL). In some embodiments, the subject has received priorsystemic treatment. In some embodiments, the non-Hodgkin lymphoma isanaplastic large cell lymphoma (ALCL). In some embodiments, theanaplastic large cell lymphoma (ALCL) is a systemic anaplastic largecell lymphoma (sALCL).

In some embodiments, the lymphoma is a Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject relapsed afterautologous stem cell transplant. In some embodiments, the subjectrelapsed after first-line treatment and the subject is ineligible forautologous stem cell transplant. In some embodiments, the subject hasnot been previously treated for the Hodgkin lymphoma. In someembodiments, the Hodgkin lymphoma is classical Hodgkin lymphoma (cHL).In some embodiments, the classical Hodgkin lymphoma (cHL) is advancedcHL. In some embodiments, the subject has been previously treated forcHL. In some embodiments, the subject has not been previously treatedfor cHL.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents capable of modulating the immuneresponse. In some embodiments, the one or more additional therapeuticagents is not an antibody or antigen-binding fragment thereof. In someembodiments, the one or more additional therapeutic agents is anantibody or antigen-binding fragment thereof.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents. In some embodiments, the one or moreadditional therapeutic agents is a chemotherapy regimen consistingessentially of doxorubicin, vinblastine, and dacarbazine (AVD). In someembodiments, the one or more additional therapeutic agents is achemotherapy regimen consisting essentially of Cyclophosphamide,Doxorubicin, and Prednisone (CHP). In some embodiments, the one or moreadditional therapeutic agents is an alkylating agent, an anthracycline,an antibiotic, an antifolate, an antimetabolite, an antitubulin agent,an auristatin, a chemotherapy sensitizer, a DNA minor groove binder, aDNA replication inhibitor, a duocarmycin, an etoposide, a fluorinatedpyrimidine, a lexitropsin, a nitrosourea, a platinol, a purineantimetabolite, a puromycin, a radiation sensitizer, a steroid, ataxane, a topoisomerase inhibitor, and/or a vinca alkaloid. In someembodiments, the one or more additional therapeutic agents is selectedfrom the group consisting of adriamycin, an androgen, anthramycin (AMC),asparaginase, 5-azacytidine, azathioprine, bleomycin, busulfan,buthionine sulfoximine, camptothecin, carboplatin, carmustine (BSNU),CC-1065, chlorambucil, cisplatin, colchicine, cyclophosphamide,cytarabine, cytidine arabinoside, cytochalasin B, dacarbazine,dactinomycin (formerly actinomycin), daunorubicin, decarbazine,docetaxel, doxorubicin, an estrogen, 5-fluordeoxyuridine,5-fluorouracil, gramicidin D, hydroxydaunorubicin, hydroxyurea,idarubicin, ifosfamide, irinotecan, lomustine (CCNU), mechlorethamine,melphalan, 6-mercaptopurine, methotrexate, mithramycin, mitomycin C,mitoxantrone, nitroimidazole, paclitaxel, plicamycin, prednisone,prednisolone, procarbizine, streptozotocin, tenoposide, 6-thioguanine,thioTEPA, topotecan, vinblastine, vincristine, vinorelbine, VP-16 andVM-26. In some embodiments, the one or more additional therapeuticagents is an antibody or antigen-binding fragment thereof.

In some embodiments, the subject has cHL that has not been previouslytreated and the one or more additional therapeutic agents areadriamycin, dacarabazine and vinblastine. In some embodiments, the cHLis advanced cHL.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and the one or more additional therapeuticagents are cyclophosphamide, hydroxydaunorubicin and prednisone.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and the one or more additional therapeuticagents are cyclophosphamide, hydroxydaunorubicin and prednisolone.

In some embodiments, the method further comprises treating the subjectwith irradiation.

C. Methods of Treatment

In some embodiments, the present disclosure is directed to a method fortreating a tumor or a subject afflicted with a tumor comprisingadministering to the subject a therapeutically effective amount of anantibody-drug conjugate, wherein the antibody-drug conjugate comprisesan anti-CD30 antibody or an antigen-binding fragment thereof conjugatedto a monomethyl auristatin (“anti-CD30 antibody-drug conjugate”). Insome embodiments, the method of treating cancer in a subject comprisesadministering to the subject an antibody-drug conjugate, wherein theactivity of CD30⁺ T regulatory (Treg) is decreased followingadministration of the antibody-drug conjugate. In some embodiments, themethod of treating cancer in a subject comprises administering to thesubject an antibody-drug conjugate, wherein the antibody-drug conjugatecomprises an anti-CD30 antibody or an antigen-binding portion thereofconjugated to a monomethyl auristatin, wherein the ratio of CD8⁺ T cellsto CD30⁺ T regulatory (Treg) cells in the subject is increased followingadministration of the antibody drug conjugate. In some embodiments, themethod of treating cancer in a subject comprises administering to thesubject an antibody-drug conjugate, wherein the antibody-drug conjugatecomprises an anti-CD30 antibody or an antigen-binding portion thereofconjugated to a monomethyl auristatin, wherein the immune response ismodulated following administration of the antibody-drug conjugate,wherein the modulation comprises increasing the ratio of CD8⁺ T cells toCD30⁺ T regulatory (Treg) cells in the subject.

In some embodiments, the tumor is derived from a Hodgkin lymphoma (HL),a non-Hodgkin lymphoma (NHL), or a combination thereof. In certainembodiments, the subject has received one, two, three, four, five ormore prior cancer treatments. In other embodiments, the subject istreatment-naïve. In some embodiments, the subject has progressed onother cancer treatments. In some embodiments, the subject has received aprevious cancer treatment and either did not respond or relapsed afterthe previous treatment. In some embodiments, the subject relapsed afterprevious cancer treatment and is ineligible for autologous stem celltransplant. In some embodiments, the subject relapsed after autologousstem cell transplant. In some embodiments, the tumor has reoccurred. Insome embodiments, the tumor is metastatic. In other embodiments, thetumor is not metastatic.

In certain embodiments, the tumor is derived from an HL (e.g., a tumorcomprising an HL). In certain embodiments, the subject has not beenpreviously treated for the HL. In certain embodiments, the subject hasbeen previously treated for the Hodgkin lymphoma and the subject did notrespond to treatment or relapsed after first-line treatment. In certainembodiments, the subject relapsed after first-line treatment and thesubject is ineligible for autologous stem cell transplant. In certainembodiments, the subject relapsed after autologous stem cell transplant.In certain embodiments, the HL is a classical HL (cHL; e.g., a nodularsclerosing HL, a mixed cellularity HL, a lymphocyte rich HL, or alymphocyte depleted HL). In other embodiments, the HL is a nodularlymphocyte predominant type HL. In certain embodiments, the subject hasnot been previously treated for the cHL. In certain embodiments, thesubject has not been previously treated for the cHL. In certainembodiments, the cHL is advanced cHL. In certain embodiments, thesubject has not been previously treated for the advanced cHL. In certainembodiments, the subject has not been previously treated for theadvanced cHL.

In other embodiments, the tumor is derived from a NHL. In someembodiments, the tumor comprises an NHL. In certain embodiments, the NHLis a relapsed or refractory NHL. In certain embodiments, the NHL has notbeen previously treated. In certain embodiments, the subject has notbeen previously treated for the NHL. In certain embodiments, the subjecthas been previously treated for the NHL and the subject did not respondto treatment or relapsed after first-line treatment. In someembodiments, the NHL is a B-cell lymphoma, e.g., a diffuse large B-celllymphoma (DLBCL), a follicular lymphoma (FL), a Burkitt lymphoma, animmunoblastic large cell lymphoma, a precursor B-lymphoblastic lymphoma,a mantle cell lymphoma, or any combination thereof. In some embodiments,the NHL is a T-cell lymphoma, e.g., a cutaneous T-cell lymphoma (CTCL),a peripheral T-cell lymphoma (PTCL), a mycosis fungoides, an anaplasticlarge cell lymphoma, a precursor T-lymphoblastic lymphoma, or anycombination thereof. In certain embodiments, the NHL is selected from aDLBCL, a PTCL, a CTCL, and any combination thereof. In certainembodiments, the NHL is a CTCL that is a relapsed or refractory CTCL. Incertain embodiments, the T-cell lymphoma is a mature T-cell lymphoma. Incertain embodiments, the subject has not been previously treated for themature T-cell lymphoma.

In some embodiments, the method of treating cancer in a subjectcomprises administering to the subject an antibody drug conjugate,wherein the antibody-drug conjugate comprises an anti-CD30 antibody oran antigen-binding portion thereof conjugated to a monomethylauristatin, wherein the activity of CD30⁺ T regulatory (Treg) isdecreased following administration of the antibody-drug conjugate. Insome embodiments, decreasing the activity of CD30⁺ Treg cells comprisesa decrease in the number of CD30⁺ Treg cells. In some embodiments, thenumber of CD30⁺ Treg cells is decreased relative to one or more othertypes of CD4⁺ T cells. In some embodiments, the one or more other typesof CD4⁺ T cells comprise Th1 cells, Th2 cells or Th17 cells. In someembodiments, the one or more other types of CD4⁺ T cells comprise Th1CD30⁺ cells, Th2 CD30⁺ cells or Th17 CD30⁺ cells. In some embodiments,the number of CD30⁺ Treg cells is decreased relative to the number ofCD30⁺ Treg cells in the subject prior to administration of theantibody-drug conjugate. In some embodiments, the number of CD30⁺ Tregcells is decreased relative to a subject who has not been treated withthe antibody-drug conjugate. In some embodiments, decreasing theactivity of CD30⁺ Treg cells comprises a decrease in the function ofCD30⁺ Treg cells. In some embodiments, the decrease in the function ofCD30⁺ Treg cells is relative to the function of CD30⁺ Treg cells in asubject prior to administration of the antibody-drug conjugate. In someembodiments, the decrease in the function of CD30⁺ Treg cells isrelative to the function of CD30⁺ Treg cells in a subject who has notbeen treated with the antibody-drug conjugate.

In some embodiments, the CD30⁺ Treg cells are CD30⁺ inducible Tregulatory (iTreg) cells or CD30⁺ peripheral T regulatory (pTreg) cells.

In some embodiments, the monomethyl auristatin is monomethyl auristatinE (MMAE). In some embodiments, the monomethyl auristatin is monomethylauristatin F (MMAF).

In some embodiments, the anti-CD30 antibody is monoclonal anti-CD30antibody AC10. In some embodiments, the anti-CD30 antibody is cAC10. Insome embodiments, the antibody-drug conjugate is brentuximab vedotin.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region and a light chain variable region, wherein the heavychain variable region comprises:

-   -   (i) a CDR-H1 comprising the amino acid sequence of SEQ ID NO: 1;    -   (ii) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:        2; and    -   (iii) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:        3; and        wherein the light chain variable region comprises:    -   (i) a CDR-L1 comprising the amino acid sequence of SEQ ID NO: 4;    -   (ii) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:        5; and    -   (iii) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:        6.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region comprising the amino acid sequence of SEQ ID NO: 7 and alight chain variable region comprising the amino acid sequence of SEQ IDNO: 8.

In some embodiments, the antibody-drug conjugate further comprises alinker between the anti-CD30 antibody or antigen-binding portion thereofand the monomethyl auristatin. In some embodiments, the linker isselected from the group consisting of a cleavable linker and anon-cleavable linker. In some embodiments, the linker is a cleavablepeptide linker. In some embodiments, the cleavable peptide linker has aformula: -MC-vc-PAB-. In some embodiments, the linker is a non-cleavablelinker having a formula: -MC-.

In some embodiments, the subject has been previously treated for thecancer. In some embodiments, the subject did not respond to treatment orrelapsed after first-line treatment. In some embodiments, the subjecthas not previously been treated for the cancer.

In some embodiments, the cancer is a lymphoma. In some embodiments, thelymphoma is a T-cell lymphoma. In some embodiments, the lymphoma is aB-cell lymphoma.

In some embodiments, the lymphoma is a non-Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the non-Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject has not beenpreviously treated for the non-Hodgkin lymphoma. In some embodiments,the non-Hodgkin lymphoma is a mature T-cell lymphoma. In someembodiments, the non-Hodgkin lymphoma is diffuse large B-cell lymphoma(DLBCL), peripheral T-cell lymphoma (PTCL), anaplastic large celllymphoma (ALCL) or cutaneous T-cell lymphoma (CTCL). In someembodiments, the non-Hodgkin lymphoma is cutaneous T-cell lymphoma(CTCL). In some embodiments, the cutaneous T-cell lymphoma (CTCL) is amycosis fungoides (MF). In some embodiments, the mycosis fungoides is aCD30-positive mycosis fungoides (MF). In some embodiments, the cutaneousT-cell lymphoma (CTCL) is a primary cutaneous anaplastic large celllymphoma (pcALCL). In some embodiments, the subject has received priorsystemic treatment. In some embodiments, the non-Hodgkin lymphoma isanaplastic large cell lymphoma (ALCL). In some embodiments, theanaplastic large cell lymphoma (ALCL) is a systemic anaplastic largecell lymphoma (sALCL)

In some embodiments, the lymphoma is a Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject relapsed afterautologous stem cell transplant. In some embodiments, the subjectrelapsed after first-line treatment and the subject is ineligible forautologous stem cell transplant. In some embodiments, the subject hasnot been previously treated for the Hodgkin lymphoma. In someembodiments, the Hodgkin lymphoma is classical Hodgkin lymphoma (cHL).In some embodiments, the classical Hodgkin lymphoma (cHL) is advancedcHL. In some embodiments, the subject has been previously treated forcHL. In some embodiments, the subject has not been previously treatedfor cHL.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents capable of modulating the immuneresponse. In some embodiments, the one or more additional therapeuticagents is not an antibody or antigen-binding fragment thereof. In someembodiments, the one or more additional therapeutic agents is anantibody or antigen-binding fragment thereof.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents. In some embodiments, the one or moreadditional therapeutic agents is a chemotherapy regimen consistingessentially of doxorubicin, vinblastine, and dacarbazine (AVD). In someembodiments, the one or more additional therapeutic agents is achemotherapy regimen consisting essentially of Cyclophosphamide,Doxorubicin, and Prednisone (CHP). In some embodiments, the one or moreadditional therapeutic agents is an alkylating agent, an anthracycline,an antibiotic, an antifolate, an antimetabolite, an antitubulin agent,an auristatin, a chemotherapy sensitizer, a DNA minor groove binder, aDNA replication inhibitor, a duocarmycin, an etoposide, a fluorinatedpyrimidine, a lexitropsin, a nitrosourea, a platinol, a purineantimetabolite, a puromycin, a radiation sensitizer, a steroid, ataxane, a topoisomerase inhibitor, and/or a vinca alkaloid. In someembodiments, the one or more additional therapeutic agents is selectedfrom the group consisting of adriamycin, an androgen, anthramycin (AMC),asparaginase, 5-azacytidine, azathioprine, bleomycin, busulfan,buthionine sulfoximine, camptothecin, carboplatin, carmustine (BSNU),CC-1065, chlorambucil, cisplatin, colchicine, cyclophosphamide,cytarabine, cytidine arabinoside, cytochalasin B, dacarbazine,dactinomycin (formerly actinomycin), daunorubicin, decarbazine,docetaxel, doxorubicin, an estrogen, 5-fluordeoxyuridine,5-fluorouracil, gramicidin D, hydroxydaunorubicin, hydroxyurea,idarubicin, ifosfamide, irinotecan, lomustine (CCNU), mechlorethamine,melphalan, 6-mercaptopurine, methotrexate, mithramycin, mitomycin C,mitoxantrone, nitroimidazole, paclitaxel, plicamycin, prednisone,prednisolone, procarbizine, streptozotocin, tenoposide, 6-thioguanine,thioTEPA, topotecan, vinblastine, vincristine, vinorelbine, VP-16 andVM-26. In some embodiments, the one or more additional therapeuticagents is an antibody or antigen-binding fragment thereof.

In some embodiments, the subject has cHL that has not been previouslytreated and wherein the one or more additional therapeutic agents areadriamycin, dacarabazine and vinblastine. In some embodiments, the cHLis advanced cHL.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and wherein the one or more additionaltherapeutic agents are cyclophosphamide, hydroxydaunorubicin andprednisone.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and wherein the one or more additionaltherapeutic agents are cyclophosphamide, hydroxydaunorubicin andprednisolone.

In some embodiments, the method further comprises treating the subjectwith irradiation.

In some embodiments, the method of treating cancer in a subjectcomprises administering to the subject an antibody-drug conjugate,wherein the antibody-drug conjugate comprises an anti-CD30 antibody oran antigen-binding portion thereof conjugated to a monomethylauristatin, wherein the ratio of CD8⁺ T cells to CD30⁺ T regulatory(Treg) cells in the subject is increased following administration of theantibody drug conjugate. In some embodiments, the ratio of CD8⁺ T cellsto CD30⁺ Treg cells is increased relative to the ratio of CD8⁺ T cellsto CD30⁺ Treg cells in the subject prior to the administration of theantibody-drug conjugate. In some embodiments, the ratio of CD8⁺ T cellsto CD30⁺ Treg cells is increased relative to the ratio of CD8⁺ T cellsto CD30⁺ Treg cells in a subject who has not been treated with theantibody-drug conjugate.

In some embodiments, the CD30⁺ Treg cells are CD30⁺ inducible Tregulatory (iTreg) cells or CD30⁺ peripheral T regulatory (pTreg) cells.

In some embodiments, the monomethyl auristatin is monomethyl auristatinE (MMAE). In some embodiments, the monomethyl auristatin is monomethylauristatin F (MMAF).

In some embodiments, the anti-CD30 antibody is monoclonal anti-CD30antibody AC10. In some embodiments, the anti-CD30 antibody is cAC10. Insome embodiments, the antibody-drug conjugate is brentuximab vedotin.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region and a light chain variable region, wherein the heavychain variable region comprises:

-   -   (i) a CDR-H1 comprising the amino acid sequence of SEQ ID NO: 1;    -   (ii) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:        2; and    -   (iii) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:        3; and        wherein the light chain variable region comprises:    -   (i) a CDR-L1 comprising the amino acid sequence of SEQ ID NO: 4;    -   (ii) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:        5; and    -   (iii) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:        6.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region comprising the amino acid sequence of SEQ ID NO: 7 and alight chain variable region comprising the amino acid sequence of SEQ IDNO: 8.

In some embodiments, the antibody-drug conjugate further comprises alinker between the anti-CD30 antibody or antigen-binding portion thereofand the monomethyl auristatin. In some embodiments, the linker isselected from the group consisting of a cleavable linker and anon-cleavable linker. In some embodiments, the linker is a cleavablepeptide linker. In some embodiments, the cleavable peptide linker has aformula: -MC-vc-PAB-. In some embodiments, the linker is a non-cleavablelinker having a formula: -MC-.

In some embodiments, the subject has been previously treated for thecancer. In some embodiments, the subject did not respond to treatment orrelapsed after first-line treatment. In some embodiments, the subjecthas not previously been treated for the cancer.

In some embodiments, the cancer is a lymphoma. In some embodiments, thelymphoma is a T-cell lymphoma. In some embodiments, the lymphoma is aB-cell lymphoma.

In some embodiments, the lymphoma is a non-Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the non-Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject has not beenpreviously treated for the non-Hodgkin lymphoma. In some embodiments,the non-Hodgkin lymphoma is a mature T-cell lymphoma. In someembodiments, the non-Hodgkin lymphoma is diffuse large B-cell lymphoma(DLBCL), peripheral T-cell lymphoma (PTCL), anaplastic large celllymphoma (ALCL) or cutaneous T-cell lymphoma (CTCL). In someembodiments, the non-Hodgkin lymphoma is cutaneous T-cell lymphoma(CTCL). In some embodiments, the cutaneous T-cell lymphoma (CTCL) is amycosis fungoides (MF). In some embodiments, the mycosis fungoides is aCD30-positive mycosis fungoides (MF). In some embodiments, the cutaneousT-cell lymphoma (CTCL) is a primary cutaneous anaplastic large celllymphoma (pcALCL). In some embodiments, the subject has received priorsystemic treatment. In some embodiments, the non-Hodgkin lymphoma isanaplastic large cell lymphoma (ALCL). In some embodiments, theanaplastic large cell lymphoma (ALCL) is a systemic anaplastic largecell lymphoma (sALCL)

In some embodiments, the lymphoma is a Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject relapsed afterautologous stem cell transplant. In some embodiments, the subjectrelapsed after first-line treatment and the subject is ineligible forautologous stem cell transplant. In some embodiments, the subject hasnot been previously treated for the Hodgkin lymphoma. In someembodiments, the Hodgkin lymphoma is classical Hodgkin lymphoma (cHL).In some embodiments, the classical Hodgkin lymphoma (cHL) is advancedcHL. In some embodiments, the subject has been previously treated forcHL. In some embodiments, the subject has not been previously treatedfor cHL.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents capable of modulating the immuneresponse. In some embodiments, the one or more additional therapeuticagents is not an antibody or antigen-binding fragment thereof. In someembodiments, the one or more additional therapeutic agents is anantibody or antigen-binding fragment thereof.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents. In some embodiments, the one or moreadditional therapeutic agents is a chemotherapy regimen consistingessentially of doxorubicin, vinblastine, and dacarbazine (AVD). In someembodiments, the one or more additional therapeutic agents is achemotherapy regimen consisting essentially of Cyclophosphamide,Doxorubicin, and Prednisone (CHP). In some embodiments, the one or moreadditional therapeutic agents is an alkylating agent, an anthracycline,an antibiotic, an antifolate, an antimetabolite, an antitubulin agent,an auristatin, a chemotherapy sensitizer, a DNA minor groove binder, aDNA replication inhibitor, a duocarmycin, an etoposide, a fluorinatedpyrimidine, a lexitropsin, a nitrosourea, a platinol, a purineantimetabolite, a puromycin, a radiation sensitizer, a steroid, ataxane, a topoisomerase inhibitor, and/or a vinca alkaloid. In someembodiments, the one or more additional therapeutic agents is selectedfrom the group consisting of adriamycin, an androgen, anthramycin (AMC),asparaginase, 5-azacytidine, azathioprine, bleomycin, busulfan,buthionine sulfoximine, camptothecin, carboplatin, carmustine (BSNU),CC-1065, chlorambucil, cisplatin, colchicine, cyclophosphamide,cytarabine, cytidine arabinoside, cytochalasin B, dacarbazine,dactinomycin (formerly actinomycin), daunorubicin, decarbazine,docetaxel, doxorubicin, an estrogen, 5-fluordeoxyuridine,5-fluorouracil, gramicidin D, hydroxydaunorubicin, hydroxyurea,idarubicin, ifosfamide, irinotecan, lomustine (CCNU), mechlorethamine,melphalan, 6-mercaptopurine, methotrexate, mithramycin, mitomycin C,mitoxantrone, nitroimidazole, paclitaxel, plicamycin, prednisone,prednisolone, procarbizine, streptozotocin, tenoposide, 6-thioguanine,thioTEPA, topotecan, vinblastine, vincristine, vinorelbine, VP-16 andVM-26. In some embodiments, the one or more additional therapeuticagents is an antibody or antigen-binding fragment thereof.

In some embodiments, the subject has cHL that has not been previouslytreated and wherein the one or more additional therapeutic agents areadriamycin, dacarabazine and vinblastine. In some embodiments, the cHLis advanced cHL.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and wherein the one or more additionaltherapeutic agents are cyclophosphamide, hydroxydaunorubicin andprednisone.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and wherein the one or more additionaltherapeutic agents are cyclophosphamide, hydroxydaunorubicin andprednisolone.

In some embodiments, the method further comprises treating the subjectwith irradiation.

In some embodiments, the method of treating cancer in a subjectcomprising administering to the subject an antibody-drug conjugate,wherein the antibody-drug conjugate comprises an anti-CD30 antibody oran antigen-binding portion thereof conjugated to a monomethylauristatin, wherein the immune response is modulated followingadministration of the antibody-drug conjugate, wherein the modulationcomprises increasing the ratio of CD8⁺ T cells to CD30⁺ T regulatory(Treg) cells in the subject. In some embodiments, the ratio of CD8⁺ Tcells to CD30⁺ Treg cells is increased relative to the ratio of CD8⁺ Tcells to CD30⁺ Treg cells in the subject prior to the administration ofthe antibody-drug conjugate. In some embodiments, the ratio of CD8⁺ Tcells to CD30⁺ Treg cells is increased relative to the ratio of CD8⁺ Tcells to CD30⁺ Treg cells in a subject who has not been treated with theantibody-drug conjugate.

In some embodiments, the CD30⁺ Treg cells are CD30⁺ inducible Tregulatory (iTreg) cells or CD30⁺ peripheral T regulatory (pTreg) cells.

In some embodiments, the monomethyl auristatin is monomethyl auristatinE (MMAE). In some embodiments, the monomethyl auristatin is monomethylauristatin F (MMAF).

In some embodiments, the anti-CD30 antibody is monoclonal anti-CD30antibody AC10. In some embodiments, the anti-CD30 antibody is cAC10. Insome embodiments, the antibody-drug conjugate is brentuximab vedotin.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region and a light chain variable region, wherein the heavychain variable region comprises:

-   -   (i) a CDR-H1 comprising the amino acid sequence of SEQ ID NO: 1;    -   (ii) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:        2; and    -   (iii) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:        3; and        wherein the light chain variable region comprises:    -   (i) a CDR-L1 comprising the amino acid sequence of SEQ ID NO: 4;    -   (ii) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:        5; and    -   (iii) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:        6.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region comprising the amino acid sequence of SEQ ID NO: 7 and alight chain variable region comprising the amino acid sequence of SEQ IDNO: 8.

In some embodiments, the antibody-drug conjugate further comprises alinker between the anti-CD30 antibody or antigen-binding portion thereofand the monomethyl auristatin. In some embodiments, the linker isselected from the group consisting of a cleavable linker and anon-cleavable linker. In some embodiments, the linker is a cleavablepeptide linker. In some embodiments, the cleavable peptide linker has aformula: -MC-vc-PAB-. In some embodiments, the linker is a non-cleavablelinker having a formula: -MC-.

In some embodiments, the subject has been previously treated for thecancer. In some embodiments, the subject did not respond to treatment orrelapsed after first-line treatment. In some embodiments, the subjecthas not previously been treated for the cancer.

In some embodiments, the cancer is a lymphoma. In some embodiments, thelymphoma is a T-cell lymphoma. In some embodiments, the lymphoma is aB-cell lymphoma.

In some embodiments, the lymphoma is a non-Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the non-Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject has not beenpreviously treated for the non-Hodgkin lymphoma. In some embodiments,the non-Hodgkin lymphoma is a mature T-cell lymphoma. In someembodiments, the non-Hodgkin lymphoma is diffuse large B-cell lymphoma(DLBCL), peripheral T-cell lymphoma (PTCL), anaplastic large celllymphoma (ALCL) or cutaneous T-cell lymphoma (CTCL). In someembodiments, the non-Hodgkin lymphoma is cutaneous T-cell lymphoma(CTCL). In some embodiments, the cutaneous T-cell lymphoma (CTCL) is amycosis fungoides (MF). In some embodiments, the mycosis fungoides is aCD30-positive mycosis fungoides (MF). In some embodiments, the cutaneousT-cell lymphoma (CTCL) is a primary cutaneous anaplastic large celllymphoma (pcALCL). In some embodiments, the subject has received priorsystemic treatment. In some embodiments, the non-Hodgkin lymphoma isanaplastic large cell lymphoma (ALCL). In some embodiments, theanaplastic large cell lymphoma (ALCL) is a systemic anaplastic largecell lymphoma (sALCL).

In some embodiments, the lymphoma is a Hodgkin lymphoma. In someembodiments, the subject has been previously treated for the Hodgkinlymphoma and the subject did not respond to treatment or relapsed afterfirst-line treatment. In some embodiments, the subject relapsed afterautologous stem cell transplant. In some embodiments, the subjectrelapsed after first-line treatment and the subject is ineligible forautologous stem cell transplant. In some embodiments, the subject hasnot been previously treated for the Hodgkin lymphoma. In someembodiments, the Hodgkin lymphoma is classical Hodgkin lymphoma (cHL).In some embodiments, the classical Hodgkin lymphoma (cHL) is advancedcHL. In some embodiments, the subject has been previously treated forcHL. In some embodiments, the subject has not been previously treatedfor cHL.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents capable of modulating the immuneresponse. In some embodiments, the one or more additional therapeuticagents is not an antibody or antigen-binding fragment thereof. In someembodiments, the one or more additional therapeutic agents is anantibody or antigen-binding fragment thereof.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents. In some embodiments, the one or moreadditional therapeutic agents is a chemotherapy regimen consistingessentially of doxorubicin, vinblastine, and dacarbazine (AVD). In someembodiments, the one or more additional therapeutic agents is achemotherapy regimen consisting essentially of Cyclophosphamide,Doxorubicin, and Prednisone (CHP). In some embodiments, the one or moreadditional therapeutic agents is an alkylating agent, an anthracycline,an antibiotic, an antifolate, an antimetabolite, an antitubulin agent,an auristatin, a chemotherapy sensitizer, a DNA minor groove binder, aDNA replication inhibitor, a duocarmycin, an etoposide, a fluorinatedpyrimidine, a lexitropsin, a nitrosourea, a platinol, a purineantimetabolite, a puromycin, a radiation sensitizer, a steroid, ataxane, a topoisomerase inhibitor, and/or a vinca alkaloid. In someembodiments, the one or more additional therapeutic agents is selectedfrom the group consisting of adriamycin, an androgen, anthramycin (AMC),asparaginase, 5-azacytidine, azathioprine, bleomycin, busulfan,buthionine sulfoximine, camptothecin, carboplatin, carmustine (BSNU),CC-1065, chlorambucil, cisplatin, colchicine, cyclophosphamide,cytarabine, cytidine arabinoside, cytochalasin B, dacarbazine,dactinomycin (formerly actinomycin), daunorubicin, decarbazine,docetaxel, doxorubicin, an estrogen, 5-fluordeoxyuridine,5-fluorouracil, gramicidin D, hydroxydaunorubicin, hydroxyurea,idarubicin, ifosfamide, irinotecan, lomustine (CCNU), mechlorethamine,melphalan, 6-mercaptopurine, methotrexate, mithramycin, mitomycin C,mitoxantrone, nitroimidazole, paclitaxel, plicamycin, prednisone,prednisolone, procarbizine, streptozotocin, tenoposide, 6-thioguanine,thioTEPA, topotecan, vinblastine, vincristine, vinorelbine, VP-16 andVM-26. In some embodiments, the one or more additional therapeuticagents is an antibody or antigen-binding fragment thereof.

In some embodiments, the subject has cHL that has not been previouslytreated and wherein the one or more additional therapeutic agents areadriamycin, dacarabazine and vinblastine. In some embodiments, the cHLis advanced cHL.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and wherein the one or more additionaltherapeutic agents are cyclophosphamide, hydroxydaunorubicin andprednisone.

In some embodiments, the subject has a mature T-cell lymphoma that hasnot been previously treated and wherein the one or more additionaltherapeutic agents are cyclophosphamide, hydroxydaunorubicin andprednisolone.

In some embodiments, the method further comprises treating the subjectwith irradiation.

In some embodiments, the method of treating cancer is a method oftreating cutaneous T-cell lymphoma (CTCL) in a subject comprisingadministering to the subject an antibody drug conjugate, wherein theantibody drug conjugate comprises an anti-CD30 antibody or anantigen-binding portion thereof conjugated to a monomethyl auristatin.In some embodiments, the ratio of CD8⁺ T cells to CD30⁺ T regulatory(Treg) in the subject is increased following administration of theantibody drug conjugate. In some embodiments, the immune response ismodulated following administration of the antibody drug conjugate,wherein the modulation comprises increasing the ratio of CD8⁺ T cells toCD30⁺ T regulatory (Treg). In some embodiments, the ratio of CD8⁺ Tcells to CD30⁺ Treg cells is increased relative to the ratio of CD8⁺ Tcells to CD30⁺ Treg cells in the subject prior to the administration ofthe antibody-drug conjugate. In some embodiments, the ratio of CD8⁺ Tcells to CD30⁺ Treg cells is increased relative to the ratio of CD8⁺ Tcells to CD30⁺ Treg cells in a subject who has not been treated with theantibody-drug conjugate. In some embodiments, the subject did notrespond to treatment for the CTCL or relapsed after first-line treatmentfor the CTCL. In some embodiments, the subject has not been previouslytreated for the CTCL.

In some embodiments, the CD30⁺ Treg cells are CD30⁺ inducible Tregulatory (iTreg) cells or CD30⁺ peripheral T regulatory (pTreg) cells.

In some embodiments, the monomethyl auristatin is monomethyl auristatinE (MMAE). In some embodiments, the monomethyl auristatin is monomethylauristatin F (MMAF).

In some embodiments, the anti-CD30 antibody is monoclonal anti-CD30antibody AC10. In some embodiments, the anti-CD30 antibody is cAC10. Insome embodiments, the antibody-drug conjugate is brentuximab vedotin.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region and a light chain variable region, wherein the heavychain variable region comprises:

-   -   (i) a CDR-H1 comprising the amino acid sequence of SEQ ID NO: 1;    -   (ii) a CDR-H2 comprising the amino acid sequence of SEQ ID NO:        2; and    -   (iii) a CDR-H3 comprising the amino acid sequence of SEQ ID NO:        3; and        wherein the light chain variable region comprises:    -   (i) a CDR-L1 comprising the amino acid sequence of SEQ ID NO: 4;    -   (ii) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:        5; and    -   (iii) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:        6.

In some embodiments, the anti-CD30 antibody comprises a heavy chainvariable region comprising the amino acid sequence of SEQ ID NO: 7 and alight chain variable region comprising the amino acid sequence of SEQ IDNO: 8.

In some embodiments, the antibody-drug conjugate further comprises alinker between the anti-CD30 antibody or antigen-binding portion thereofand the monomethyl auristatin. In some embodiments, the linker isselected from the group consisting of a cleavable linker and anon-cleavable linker. In some embodiments, the linker is a cleavablepeptide linker. In some embodiments, the cleavable peptide linker has aformula: -MC-vc-PAB-. In some embodiments, the linker is a non-cleavablelinker having a formula: -MC-.

In some embodiments, the subject has been previously treated for thecancer. In some embodiments, the subject did not respond to treatment orrelapsed after first-line treatment. In some embodiments, the subjecthas not previously been treated for the cancer.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents capable of modulating the immuneresponse. In some embodiments, the one or more additional therapeuticagents is not an antibody or antigen-binding fragment thereof. In someembodiments, the one or more additional therapeutic agents is anantibody or antigen-binding fragment thereof.

In some embodiments, the method further comprises administering one ormore additional therapeutic agents. In some embodiments, the one or moreadditional therapeutic agents is a chemotherapy regimen consistingessentially of doxorubicin, vinblastine, and dacarbazine (AVD). In someembodiments, the one or more additional therapeutic agents is achemotherapy regimen consisting essentially of Cyclophosphamide,Doxorubicin, and Prednisone (CHP). In some embodiments, the one or moreadditional therapeutic agents is an alkylating agent, an anthracycline,an antibiotic, an antifolate, an antimetabolite, an antitubulin agent,an auristatin, a chemotherapy sensitizer, a DNA minor groove binder, aDNA replication inhibitor, a duocarmycin, an etoposide, a fluorinatedpyrimidine, a lexitropsin, a nitrosourea, a platinol, a purineantimetabolite, a puromycin, a radiation sensitizer, a steroid, ataxane, a topoisomerase inhibitor, and/or a vinca alkaloid. In someembodiments, the one or more additional therapeutic agents is selectedfrom the group consisting of adriamycin, an androgen, anthramycin (AMC),asparaginase, 5-azacytidine, azathioprine, bleomycin, busulfan,buthionine sulfoximine, camptothecin, carboplatin, carmustine (BSNU),CC-1065, chlorambucil, cisplatin, colchicine, cyclophosphamide,cytarabine, cytidine arabinoside, cytochalasin B, dacarbazine,dactinomycin (formerly actinomycin), daunorubicin, decarbazine,docetaxel, doxorubicin, an estrogen, 5-fluordeoxyuridine,5-fluorouracil, gramicidin D, hydroxydaunorubicin, hydroxyurea,idarubicin, ifosfamide, irinotecan, lomustine (CCNU), mechlorethamine,melphalan, 6-mercaptopurine, methotrexate, mithramycin, mitomycin C,mitoxantrone, nitroimidazole, paclitaxel, plicamycin, prednisone,prednisolone, procarbizine, streptozotocin, tenoposide, 6-thioguanine,thioTEPA, topotecan, vinblastine, vincristine, vinorelbine, VP-16 andVM-26. In some embodiments, the one or more additional therapeuticagents is an antibody or antigen-binding fragment thereof.

In some embodiments, the method further comprises treating the subjectwith irradiation.

In some embodiments, the method of treating cancer is a method oftreating a non-Hodgkin lymphoma or a Hodgkin lymphoma in a subjectcomprising administering to the subject an antibody drug conjugate,wherein the antibody drug conjugate comprises an anti-CD30 antibody oran antigen-binding portion thereof conjugated to a monomethylauristatin. In some embodiments, the ratio of CD8⁺ T cells to CD30⁺ Tregulatory (Treg) in the subject is increased following administrationof the antibody drug conjugate. In some embodiments, the immune responseis modulated following administration of the antibody drug conjugate,wherein the modulation comprises increasing the ratio of CD8⁺ T cells toCD30⁺ T regulatory (Treg). In some embodiments, the subject has notpreviously been treated for the non-Hodgkin lymphoma or Hodgkinlymphoma. In some embodiments, the cancer is a non-Hodgkin lymphoma. Insome embodiments, the subject has not previously been treated for thenon-Hodgkin lymphoma. In some embodiments, non-Hodgkin lymphoma is amature T-cell lymphoma. In some embodiments the subject has not beenpreviously been treated for the mature T-cell lymphoma and the methodfurther comprises administering one or more additional therapeuticagents. In some embodiments, the one or more additional therapeuticagents comprise one or more agents selected from the group consisting ofcyclophosphamide, hydroxydaunorubicin, prednisone and prednisolone. Insome embodiments, the one or more additional therapeutic agents is achemotherapy regimen consisting essentially of cyclophosphamide,hydroxydaunorubicin and prednisone. In some embodiments, the one or moreadditional therapeutic agents is a chemotherapy regimen consistingessentially of cyclophosphamide, hydroxydaunorubicin and prednisolone.In some embodiments, the cancer is a Hodgkin lymphoma. In someembodiments, the Hodgkin lymphoma is classical Hodgkin lymphoma. In someembodiments, the subject has not been previously treated for theclassical Hodgkin lymphoma and the method further comprisesadministering one or more additional therapeutic agents. In someembodiments, the one or more additional therapeutic agents is achemotherapy regimen consisting essentially of adriamycin, vinblastineand dacarbazine. In some embodiments, the one or more additionaltherapeutic agents is a chemotherapy regimen consisting essentially ofadriamycin, vinblastine and dacarbazine.

In some embodiments, the present methods further comprise administeringone or more additional therapeutic agents capable of modulating theimmune response. In some embodiments, the one or more additionaltherapeutic agents is not an antibody or antigen-binding fragmentthereof. In some embodiments, the one or more additional therapeuticagents is an antibody or antigen-binding fragment thereof.

In some embodiments, the present methods further comprise administeringone or more additional therapeutic agents. In some embodiments, the oneor more additional therapeutic agents is an alkylating agent, ananthracycline, an antibiotic, an antifolate, an antimetabolite, anantitubulin agent, an auristatin, a chemotherapy sensitizer, a DNA minorgroove binder, a DNA replication inhibitor, a duocarmycin, an etoposide,a fluorinated pyrimidine, a lexitropsin, a nitrosourea, a platinol, apurine antimetabolite, a puromycin, a radiation sensitizer, a steroid, ataxane, a topoisomerase inhibitor, and/or a vinca alkaloid. In someembodiments, the one or more additional therapeutic agents is selectedfrom the group consisting of adriamycin, an androgen, anthramycin (AMC),asparaginase, 5-azacytidine, azathioprine, bleomycin, busulfan,buthionine sulfoximine, camptothecin, carboplatin, carmustine (BSNi),CC-1065, chlorambucil, cisplatin, colchicine, cyclophosphanide,cytarabine, cytidine arabinoside, cytochalasin B, dacarbazine,dactinomycin (formerly actinomycin), daunorubicin, decarbazine,docetaxel, doxorubicin, an estrogen, 5-fluordeoxyuridine,5-fluorouracil, gramicidin D, hydroxydaunorubicin, hydroxyurea,idarubicin, ifosfamide, irinotecan, lomustine (CCNU), mechlorethamine,melphalan, 6-mercaptopurine, methotrexate, mithramycin, mitomycin C,mitoxantrone, nitroimidazole, paclitaxel, plicamycin, prednisone,prednisolone, procarbizine, streptozotocin, tenoposide, 6-thioguanine,thioTEPA, topotecan, vinblastine, vincristine, vinorelbine, VP-16 andVM-26. In some embodiments, the one or more additional therapeuticagents are adriamycin, dacarabazine and vinblastine. In someembodiments, the one or more additional therapeutic agents arecyclophosphamide, hydroxydaunorubicin and prednisone. In someembodiments, the one or more additional therapeutic agents arecyclophosphamide, hydroxydaunorubicin and prednisolone.

In other embodiments, the present methods comprise administering aneffective amount of an anti-CD30 antibody-drug conjugate. An effectiveamount of an anti-CD30 antibody-drug conjugate can be a flat dose or aweight based dose.

In certain embodiments, the therapy of the present disclosure (e.g.,administration of an anti-CD30 antibody-drug conjugate) effectivelyincreases the duration of survival of the subject. For example, theduration of survival of the subject is increased by at least about 1month, at least about 2 months, at least about 3 months, at least about4 months, at least about 5 months, at least about 6 months, at leastabout 7 months, at least about 8 months, at least about 9 months, atleast about 10 months, at least about 11 months, or at least about 1year or more when compared to another subject treated with anothertherapy.

In certain embodiments, the therapy of the present disclosureeffectively increases the duration of progression-free survival of thesubject. For example, the progression free survival of the subject isincreased by at least about 1 month, at least about 2 months, at leastabout 3 months, at least about 4 months, at least about 5 months, atleast about 6 months, at least about 7 months, at least about 8 months,at least about 9 months, at least about 10 months, at least about 11months, or at least about 1 year when compared to another subjecttreated with another therapy

In certain embodiments, the therapy of the present disclosureeffectively increases the response rate in a group of subjects. Forexample, the response rate in a group of subjects is increased by atleast about 2%, at least about 3%, at least about 4%, at least about 5%,at least about 10%, at least about 15%, at least about 20%, at leastabout 25%, at least about 30%, at last about 35%, at least about 40%, atleast about 45%, at least about 50%, at least about 55%, at least about60%, at least about 70%, at least about 75%, at least about 80%, atleast about 85%, at least about 90%, at least about 95%, at least about99% or at least about 100% when compared to another group of subjectstreated with another therapy.

III. Compositions

In some aspects, provided herein are compositions (e.g., pharmaceuticalcompositions) comprising any of the anti-CD30 antibody-drug conjugatesdescribed herein (e.g., an anti-CD30 antibody drug conjugate that bindsto human CD30). The anti-CD30 drug-conjugates of the present disclosurecan be constituted in a composition, e.g., a pharmaceutical compositioncontaining an antibody drug-conjugate and a pharmaceutically acceptablecarrier. As used herein, a “pharmaceutically acceptable carrier”includes any and all solvents, dispersion media, coatings, antibacterialand antifungal agents, isotonic and absorption delaying agents, and thelike that are physiologically compatible. In some embodiments, thecarrier for a composition containing an antibody drug-conjugate issuitable for intravenous, intramuscular, subcutaneous, parenteral,spinal, or epidermal administration (e.g., by injection or infusion). Apharmaceutical composition of the disclosure can include one or morepharmaceutically acceptable salts, anti-oxidants, aqueous andnon-aqueous carriers, and/or adjuvants such as preservatives, wettingagents, emulsifying agents, and dispersing agents.

Therapeutic formulations are prepared for storage by mixing the activeingredient having the desired degree of purity with optionalpharmaceutically acceptable carriers, excipients or stabilizers(Remington: The Science and Practice of Pharmacy, 20th Ed., LippincottWilliams & Wiklins, Pub., Gennaro Ed., Philadelphia, Pa. 2000).Acceptable carriers, excipients, or stabilizers are nontoxic torecipients at the dosages and concentrations employed, and includebuffers, antioxidants including ascorbic acid, methionine, Vitamin E,sodium metabisulfite; preservatives, isotonicifiers, stabilizers, metalcomplexes (e.g., Zn-protein complexes); chelating agents such as EDTAand/or non-ionic surfactants.

Buffers can be used to control the pH in a range which optimizes thetherapeutic effectiveness, especially if stability is pH dependent.Buffers can be present at concentrations ranging from about 50 mM toabout 250 mM. Suitable buffering agents for use with the presentinvention include both organic and inorganic acids and salts thereof.For example, citrate, phosphate, succinate, tartrate, fumarate,gluconate, oxalate, lactate, acetate. Additionally, buffers may becomprised of histidine and trimethylamine salts such as Tris.

Preservatives can be added to prevent microbial growth, and aretypically present in a range from about 0.2%-1.0% (w/v). Suitablepreservatives for use with the present invention includeoctadecyldimethylbenzyl ammonium chloride; hexamethonium chloride;benzalkonium halides (e.g., chloride, bromide, iodide), benzethoniumchloride; thimerosal, phenol, butyl or benzyl alcohol; alkyl parabenssuch as methyl or propyl paraben; catechol; resorcinol; cyclohexanol,3-pentanol, and m-cresol.

Tonicity agents, sometimes known as “stabilizers” can be present toadjust or maintain the tonicity of liquid in a composition. When usedwith large, charged biomolecules such as proteins and antibodies, theyare often termed “stabilizers” because they can interact with thecharged groups of the amino acid side chains, thereby lessening thepotential for inter and intra-molecular interactions. Tonicity agentscan be present in any amount between about 0.1% to about 25% by weightor between about 1 to about 5% by weight, taking into account therelative amounts of the other ingredients. In some embodiments, tonicityagents include polyhydric sugar alcohols, trihydric or higher sugaralcohols, such as glycerin, erythritol, arabitol, xylitol, sorbitol andmannitol.

Additional excipients include agents which can serve as one or more ofthe following: (1) bulking agents, (2) solubility enhancers, (3)stabilizers and (4) and agents preventing denaturation or adherence tothe container wall. Such excipients include: polyhydric sugar alcohols(enumerated above); amino acids such as alanine, glycine, glutamine,asparagine, histidine, arginine, lysine, ornithine, leucine,2-phenylalanine, glutamic acid, threonine, etc.; organic sugars or sugaralcohols such as sucrose, lactose, lactitol, trehalose, stachyose,mannose, sorbose, xylose, ribose, ribitol, myoinisitose, myoinisitol,galactose, galactitol, glycerol, cyclitols (e.g., inositol),polyethylene glycol; sulfur containing reducing agents, such as urea,glutathione, thioctic acid, sodium thioglycolate, thioglycerol,α-monothioglycerol and sodium thio sulfate; low molecular weightproteins such as human serum albumin, bovine serum albumin, gelatin orother immunoglobulins; hydrophilic polymers such aspolyvinylpyrrolidone; monosaccharides (e.g., xylose, mannose, fructose,glucose; disaccharides (e.g., lactose, maltose, sucrose); trisaccharidessuch as raffinose; and polysaccharides such as dextrin or dextran.

Non-ionic surfactants or detergents (also known as “wetting agents”) canbe present to help solubilize the therapeutic agent (e.g., anti-CD30antibody drug-conjugate) as well as to protect the therapeutic protein(e.g., anti-CD30 antibody) against agitation-induced aggregation, whichalso permits the formulation to be exposed to shear surface stresswithout causing denaturation of the active therapeutic protein.Non-ionic surfactants are present in a range of about 0.05 mg/ml toabout 1.0 mg/ml or about 0.07 mg/ml to about 0.2 mg/ml. In someembodiments, non-ionic surfactants are present in a range of about0.001% to about 0.1% w/v or about 0.01% to about 0.1% w/v or about 0.01%to about 0.025% w/v.

Suitable non-ionic surfactants include polysorbates (20, 40, 60, 65, 80,etc.), polyoxamers (184, 188, etc.), PLURONIC® polyols, TRITON®,polyoxyethylene sorbitan monoethers (TWEEN®-20, TWEEN®-80, etc.),lauromacrogol 400, polyoxyl 40 stearate, polyoxyethylene hydrogenatedcastor oil 10, 50 and 60, glycerol monostearate, sucrose fatty acidester, methyl celluose and carboxymethyl cellulose. Anionic detergentsthat can be used include sodium lauryl sulfate, dioctyle sodiumsulfosuccinate and dioctyl sodium sulfonate. Cationic detergents includebenzalkonium chloride or benzethonium chloride.

In order for the formulations to be used for in vivo administration,they must be sterile. The formulation may be rendered sterile byfiltration through sterile filtration membranes. The therapeuticcompositions herein generally are placed into a container having asterile access port, for example, an intravenous solution bag or vialhaving a stopper pierceable by a hypodermic injection needle.

The route of administration is in accordance with known and acceptedmethods, such as by single or multiple bolus or infusion over a longperiod of time in a suitable manner, e.g., injection or infusion bysubcutaneous, intravenous, intraperitoneal, intramuscular,intraarterial, intralesional or intraarticular routes, topicaladministration, inhalation or by sustained release or extended-releasemeans.

The formulation herein may also contain more than one active compound asnecessary for the particular indication being treated, preferably thosewith complementary activities that do not adversely affect each other.Alternatively, or in addition, the composition may comprise analkylating agent, an anthracycline, an antibiotic, an antifolate, anantimetabolite, an antitubulin agent, an auristatin, a cheiotherapysensitizer, a DNA minor groove binder, a DNA replication inhibitor, aduocarmycin, an etoposide, a fluorinated pyrimidine, a lexitropsin, anitrosourea, a platinol, a purine antimetabolite, a puromycin, aradiation sensitizer, a steroid, a taxane, a topoisomerase inhibitor,and/or a vinca alkaloid. In some embodiments, the composition maycomprise adriamycin, an androgen, anthramycin (AMC), asparaginase,5-azacytidine, azathioprine, bleomycin, busulfan, buthioninesulfoximine, camptothecin, carboplatin, carmustine (BSNU), CC-1065,chlorambucil, cisplatin, colchicine, cyclophosphamide, cytarabine,cytidine arabinoside, cytochalasin B, dacarbazine, dactinomycin(formerly actinomycin), daunorubicin, decarbazine, docetaxel,doxorubicin, an estrogen, 5-fluordeoxyuridine, 5-fluorouracil,gramicidin D, hydroxydaunorubicin, hydroxyurea, idarubicin, ifosfamide,irinotecan, lomustine (CCNU), mechlorethamine, melphalan,6-mercaptopurine, methotrexate, mithramycin, mitomycin C, mitoxantrone,nitroimidazole, paclitaxel, plicamycin, prednisone, prednisolone,procarbizine, streptozotocin, tenoposide, 6-thioguanine, thioTEPA,topotecan, vinblastine, vincristine, vinorelbine, VP-16 or VM-26. Insome embodiments, the composition may comprise Cyclophosphamide,Doxorubicin, and Prednisone (CHP). In some embodiments, the compositionmay comprise cyclophosphamide, Doxorubicin and Prednisolone. In someembodiments, the composition may comprise doxorubicin, vinblastine, anddacarbazine (AVD). Such molecules are suitably present in combination inamounts that are effective for the purpose intended.

Dosage regimens are adjusted to provide the optimum desired response,e.g., a maximal therapeutic response and/or minimal adverse effects. Insome embodiments, the anti-CD30 antibody drug-conjugate (e.g.,brentuximab vedotin) is administered at a weight-based dose. Foradministration of an anti-CD30 antibody drug-conjugate (e.g.,brentuximab vedotin), the dosage can range from about 0.01 mg/kg toabout 20 mg/kg, about 0.05 mg/kg to about 20 mg/kg, about 0.1 mg/kg toabout 20 mg/kg, about 0.1 mg/kg to about 15 mg/kg, about 0.1 mg/kg toabout 10 mg/kg, about 0.1 mg/kg to about 5 mg/kg, about 0.1 mg/kg toabout 4 mg/kg, about 0.1 mg/kg to about 3 mg/kg, about 0.1 to about 2mg/kg, about 1 to about 10 mg/kg, about 1 to about 10 mg/kg, about 1 toabout 8 mg/kg, about 1 to about 5 mg/kg, about 1 to about 3 mg/kg, about1 to about 2 mg/kg of the subject's body weight. For example, dosagescan be about 0.05 mg/kg, about 0.1 mg/kg, about 0.2 mg/kg, about 0.3mg/kg, about 0.4 mg/kg, about 0.5 mg/kg, about 0.6 mg/kg, about 0.7mg/kg, about 0.8 mg/kg, about 0.9 mg/kg, about 1.0 mg/kg, about 1.1mg/kg, about 1.2 mg/kg, about 1.3 mg/kg, about 1.4 mg/kg, about 1.5mg/kg, about 1.6 mg/kg, about 1.7 mg/kg, about 1.8 mg/kg, about 1.9mg/kg, about 2.0 mg/kg, about 2.1 mg/kg, about 2.2 mg/kg, about 2.3mg/kg, about 2.4 mg/kg, about 2.5 mg/kg, about 2.6 mg/kg, about 2.7mg/kg, about 2.8 mg/kg, about 2.9 mg/kg, about 3 mg/kg, about 4 mg/kg,about 5 mg/kg, about 6 mg/kg, about 7 mg/kg, about 8 mg/kg, about 9mg/kg, about 10 mg/kg, about 11 mg/kg, about 12 mg/kg, about 13 mg/kg,about 14 mg/kg, about 15 mg/kg, or about 20 mg/kg of the subject's bodyweight.

In some embodiments, the dosage of the anti-CD30 antibody drug-conjugate(e.g., brentuximab vedotin) is 0.1 mg/kg body weight. In otherembodiments, the dosage of the anti-CD30 antibody drug-conjugate (e.g.,brentuximab vedotin) is 0.2 mg/kg body weight. In other embodiments, thedosage of the anti-CD30 antibody drug-conjugate (e.g., brentuximabvedotin) is 0.3 mg/kg body weight. In other embodiments, the dosage ofthe anti-CD30 antibody drug-conjugate (e.g., brentuximab vedotin) is 0.4mg/kg body weight. In other embodiments, the dosage of the anti-CD30antibody drug-conjugate (e.g., brentuximab vedotin) is 0.5 mg/kg bodyweight. In other embodiments, the dosage of the anti-CD30 antibodydrug-conjugate (e.g., brentuximab vedotin) is 0.6 mg/kg body weight. Inother embodiments, the dosage of the anti-CD30 antibody drug-conjugate(e.g., brentuximab vedotin) is 0.7 mg/kg body weight. In otherembodiments, the dosage of the anti-CD30 antibody drug-conjugate (e.g.,brentuximab vedotin) is 0.8 mg/kg body weight. In other embodiments, thedosage of the anti-CD30 antibody drug-conjugate (e.g., brentuximabvedotin) is 0.9 mg/kg body weight. In other embodiments, the dosage ofthe anti-CD30 antibody drug-conjugate (e.g., brentuximab vedotin) is 1.0mg/kg body weight. In other embodiments, the dosage of the anti-CD30antibody drug-conjugate (e.g., brentuximab vedotin) is 1.1 mg/kg bodyweight. In other embodiments, the dosage of the anti-CD30 antibodydrug-conjugate (e.g., brentuximab vedotin) is 1.2 mg/kg body weight. Inother embodiments, the dosage of the anti-CD30 antibody drug-conjugate(e.g., brentuximab vedotin) is 1.3 mg/kg body weight. In otherembodiments, the dosage of the anti-CD30 antibody drug-conjugate (e.g.,brentuximab vedotin) is 1.4 mg/kg body weight. In other embodiments, thedosage of the anti-CD30 antibody drug-conjugate (e.g., brentuximabvedotin) is 1.5 mg/kg body weight. In other embodiments, the dosage ofthe anti-CD30 antibody drug-conjugate (e.g., brentuximab vedotin) is 1.6mg/kg body weight. In other embodiments, the dosage of the anti-CD30antibody drug-conjugate (e.g., brentuximab vedotin) is 1.7 mg/kg bodyweight. In other embodiments, the dosage of the anti-CD30 antibodydrug-conjugate (e.g., brentuximab vedotin) is 1.8 mg/kg body weight. Inother embodiments, the dosage of the anti-CD30 antibody drug-conjugate(e.g., brentuximab vedotin) is 1.9 mg/kg body weight. In otherembodiments, the dosage of the anti-CD30 antibody drug-conjugate (e.g.,brentuximab vedotin) is 2.0 mg/kg body weight. In other embodiments, thedosage of the anti-CD30 antibody drug-conjugate (e.g., brentuximabvedotin) is 2.1 mg/kg body weight. In other embodiments, the dosage ofthe anti-CD30 antibody drug-conjugate (e.g., brentuximab vedotin) is 2.2mg/kg body weight. In other embodiments, the dosage of the anti-CD30antibody drug-conjugate (e.g., brentuximab vedotin) is 2.3 mg/kg bodyweight. In other embodiments, the dosage of the anti-CD30 antibodydrug-conjugate (e.g., brentuximab vedotin) is 2.4 mg/kg body weight. Inother embodiments, the dosage of the anti-CD30 antibody drug-conjugate(e.g., brentuximab vedotin) is 2.5 mg/kg body weight. In otherembodiments, the dosage of the anti-CD30 antibody drug-conjugate (e.g.,brentuximab vedotin) is about 5 mg/kg body weight. In other embodiments,the dosage of the anti-CD30 antibody drug-conjugate (e.g., brentuximabvedotin) is about 10 mg/kg body weight.

In certain embodiments, an anti-CD30 antibody drug-conjugate (e.g.,brentuximab vedotin) is administered at a flat dose. In someembodiments, the flat dose of the anti-CD30 antibody is a dose (e.g.,flat dose) of at least about 1 to about 1500 mg, at least about 10 toabout 1000 mg, such as, at least about 50 to about 800 mg, at leastabout 100 to about 600 mg, at least about 100 to about 400 mg or atleast about 100 to about 200 mg, such as at least about 1 mg, at leastabout 3 mg, at least about 5 mg, at least about 8 mg, at least about 10mg, at least about 20 mg, at least about 30 mg, at least about 40 mg, atleast about 50 mg, at least about 60 mg, at least about 70 mg, at leastabout 80 mg, at least about 90 mg, at least about 100 mg, at least about110 mg, at least about 120 mg, at least about 130 mg, at least about 140mg, at least about 150 mg, at least about 160 mg, at least about 170 mg,at least about 180 mg, at least about 190 mg, at least about 200 mg, atleast about 220 mg, at least about 240 mg, at least about 260 mg, atleast about 280 mg, at least about 300 mg, at least about 320 mg, atleast about 340 mg, at least about 360 mg, at least about 380 mg, atleast about 400 mg, at least about 420 mg, at least about 440 mg, atleast about 460 mg, at least about 480 mg, at least about 500 mg, atleast about 600 mg, at least about 700 mg, at least about 800 mg, atleast about 900 mg, at least about 1000 mg, at least about 1100 mg, atleast about 1200 mg, at least about 1300 mg, at least about 1400 mg, orat least about 1500 mg.

In certain embodiments, an anti-CD30 antibody drug-conjugate describedherein (e.g., brentuximab vedotin) is administered at a flat dose. Insome embodiments, the flat dose of the anti-CD30 antibody drug-conjugateis a dose (e.g., flat dose) of about 1 to about 1500 mg, about 10 toabout 1000 mg, such as, about 50 to about 800 mg, about 100 to about 600mg, about 100 to about 400 mg or about 100 to about 200 mg, such asabout 1 mg, about 3 mg, about 5 mg, about 8 mg, about 10 mg, about 20mg, about 30 mg, about 40 mg, about 50 mg, about 60 mg, about 70 mg,about 80 mg, about 90 mg, about 100 mg, about 110 mg, about 120 mg,about 130 mg, about 140 mg, about 150 mg, about 160 mg, about 170 mg,about 180 mg, about 190 mg, about 200 mg, about 220 mg, about 240 mg,about 260 mg, about 280 mg, about 300 mg, about 320 mg, about 340 mg,about 360 mg, about 380 mg, about 400 mg, about 420 mg, about 440 mg,about 460 mg, about 480 mg, about 500 mg, about 600 mg, about 700 mg,about 800 mg, about 900 mg, about 1000 mg, about 1100 mg, about 1200 mg,about 1300 mg, about 1400 mg, or about 1500 mg.

An exemplary dosage regimen entails administration once per week, onceabout every 2 weeks, once about every 3 weeks, once about every 4 weeks,once about a month, once about every 3-6 months or longer. In certainembodiments, the anti-CD30 antibody drug-conjugate (e.g., brentuximabvedotin) is administered once about every 3 weeks.

In some embodiments, a subtherapeutic dose of an anti-CD30 antibodydrug-conjugate (e.g., brentuximab vedotin) is used in the methodsherein. The subtherapeutic dosages of an anti-CD30 antibodydrug-conjugate (e.g., brentuximab vedotin) used in the methods hereinare higher than 0.001 mg/kg and lower than 10 mg/kg. In someembodiments, the subtherapeutic dose is about 0.001 mg/kg-about 10mg/kg, about 0.01 mg/kg-about 10 mg/kg, about 0.01 mg/kg-about 1 mg/kg,about 0.1 mg/kg-about 1 mg/kg, or about 0.001 mg/kg-about 0.1 mg/kg bodyweight. In some embodiments, the subtherapeutic dose is at least about0.001 mg/kg, at least about 0.005 mg/kg, at least about 0.01 mg/kg, atleast about 0.05 mg/kg, at least about 0.1 mg/kg, at least about 0.2mg/kg, at least about 0.3 mg/kg, at least about 0.4 mg/kg, at leastabout 0.5 mg/kg, at least about 0.6 mg/kg, at least about 0.7 mg/kg, atleast about 0.8 mg/kg, at least about 0.9 mg/kg, at least about 1 mg/kg,at least about 1.1 mg/kg, at least about 1.2 mg/kg, at least about 1.3mg/kg, at least about 1.4 mg/kg, at least about 1.5 mg/kg, at leastabout 1.6 mg/kg, or at least about 1.7 mg/kg body weight.

In some embodiments, treatment is continued as long as clinical benefitis observed or until unacceptable toxicity or disease progressionoccurs.

Dosage and frequency vary depending on the half-life of the therapeuticagent (e.g., anti-CD30 antibody drug-conjugate) in the subject. Ingeneral, human antibodies show the longest half-life, followed byhumanized antibodies, chimeric antibodies, and nonhuman antibodies. Thedosage and frequency of administration can vary depending on whether thetreatment is prophylactic or therapeutic. In prophylactic applications,a relatively low dosage is typically administered at relativelyinfrequent intervals over a long period of time. Some patients continueto receive treatment for the rest of their lives. In therapeuticapplications, a relatively high dosage at relatively short intervals issometimes required until progression of the disease is reduced orterminated, and until the patient shows partial or complete ameliorationof symptoms of disease. Thereafter, the patient can be administered aprophylactic regime.

Actual dosage levels of the active ingredients in the pharmaceuticalcompositions of the present disclosure can be varied so as to obtain anamount of the active ingredient which is effective to achieve thedesired therapeutic response for a particular patient, composition, andmode of administration, without being unduly toxic to the patient. Theselected dosage level will depend upon a variety of pharmacokineticfactors including the activity of the particular compositions of thepresent disclosure employed, the route of administration, the time ofadministration, the rate of excretion of the particular compound beingemployed, the duration of the treatment, other drugs, compounds and/ormaterials used in combination with the particular compositions employed,the age, sex, weight, condition, general health, and prior medicalhistory of the patient being treated, and like factors well known in themedical arts. A composition of the present disclosure can beadministered via one or more routes of administration using one or moreof a variety of methods well known in the art. As will be appreciated bythe skilled artisan, the route and/or mode of administration will varydepending upon the desired results.

IV. Articles of Manufacture or Kits

Also within the scope of the present disclosure provides an article ofmanufacture or kit which comprises a therapeutic agent described herein(e.g., an anti-CD30 antibody drug-conjugate). The article of manufactureor kit may further comprise instructions for use of the therapeuticagent (e.g., an anti-CD30 antibody drug-conjugate) in the methods of theinvention. An article of manufacture or kit typically includes a labelindicating the intended use of the contents of the article ofmanufacture or kit and instructions for use. The term label includes anywriting, or recorded material supplied on or with the article ofmanufacture or kit. Thus, in certain embodiments, the article ofmanufacture or kit comprises instructions for the use of an anti-CD30antibody drug-conjugate (e.g., brentuximab vedotin) in any of themethods disclosed herein such as in a method of decreasing the activityof CD30⁺ T regulatory (Treg) cells in a subject having cancer and/or ina method of increasing the ratio of CD8⁺ T cells to CD30⁺ T regulatory(Treg) cells in a subject having cancer. In some embodiments, thesubject is a human.

Ins some embodiments, provided herein is an article of manufacture orkit for treating a subject afflicted with a cancer (e.g., having acancer), the kit comprising: (a) a dosage ranging from about 0.1 mg toabout 500 mg of an anti-CD30 antibody drug-conjugate; and (b)instructions for using the anti-CD30 antibody drug-conjugate in any ofthe methods disclosed herein. In certain embodiments for treating humanpatients, the article of manufacture or kit comprises an anti-human CD30antibody drug-conjugate disclosed herein, e.g., brentuximab vedotin.

The article of manufacture or kit may further comprise a container.Suitable containers include, for example, bottles, vials (e.g., dualchamber vials), syringes (such as single or dual chamber syringes) andtest tubes. The container may be formed from a variety of materials suchas glass or plastic. The container holds the formulation.

The article of manufacture or kit may further comprise a label or apackage insert, which is on or associated with the container, mayindicate directions for reconstitution and/or use of the formulation.The label or package insert may further indicate that the formulation isuseful or intended for subcutaneous, intravenous, or other modes ofadministration in an individual. The container holding the formulationmay be a single-use vial or a multi-use vial, which allows for repeatadministrations of the reconstituted formulation. The article ofmanufacture or kit may further comprise a second container comprising asuitable diluent. The article of manufacture or kit may further includeother materials desirable from a commercial, therapeutic, and userstandpoint, including other buffers, diluents, filters, needles,syringes, and package inserts with instructions for use.

In a specific embodiment, the present invention provides kits for asingle dose-administration unit. Such kits comprise a container of anaqueous formulation of therapeutic antibody, including both single ormulti-chambered pre-filled syringes. Exemplary pre-filled syringes areavailable from Vetter GmbH, Ravensburg, Germany.

The present invention also provides an anti-CD30 antibody drug-conjugatedescribed herein that binds to CD30 (e.g., human CD30) in combinationwith one or more therapeutic agent (e.g., a second therapeutic agent)for us in any of the methods disclosed herein such as in a method ofdecreasing the activity of CD30⁺ T regulatory (Treg) cells in a subjecthaving cancer and/or in a method of increasing the ratio of CD8⁺ T cellsto CD30⁺ T regulatory (Treg) cells in a subject having cancer. In someembodiments, the article of manufacture or kit herein optionally furthercomprises a container comprising a second therapeutic medicament (e.g.,a second therapeutic agent), wherein the anti-CD30 antibodydrug-conjugate is a first medicament (e.g., a first therapeutic agent),and which article or kit further comprises instructions on the label orpackage insert for treating the individual with the second medicament,in an effective amount. In some embodiments, the kit further comprisesan alkylating agent, an anthracycline, an antibiotic, an antifolate, anantimetabolite, an antitubulin agent, an auristatin, a chemotherapysensitizer, a DNA minor groove binder, a DNA replication inhibitor, aduocarmycin, an etoposide, a fluorinated pyrimidine, a lexitropsin, anitrosourea, a platinol, a purine antinetabolite, a puronycin, aradiation sensitizer, a steroid, a taxane, a topoisomerase inhibitor ora vinca alkaloid. In some embodiments, kit further comprises a secondtherapeutic agent is selected from the group consisting of adriamycin,an androgen, anthramycin (AMC), asparaginase, 5-azacytidine,azathioprine, bleomycin, busulfan, buthionine sulfoximine, camptothecin,carboplatin, carmustine (BSNU), CC-1065, chlorambucil, cisplatin,colchicine, cyclophosphanide, cytarabine, cytidine arabinoside,cytochalasin B, dacarbazine, dactinomycin (formerly actinomycin),daunorubicin, decarbazine, docetaxel, doxorubicin, an estrogen,5-fluordeoxyuridine, 5-fluorouracil, granicidin D, hydroxydaunorubicin,hydroxyurea, idarubicin, ifosfarnide, irinotecan, loiustine (CCNU),nechloretharnine, melphalan, 6-mercaptopurine, mettotrexate,mithramycin, mitomycin C, mitoxanrone, nitroimidazole, paclitaxel,plicamycin, prednisone, prednisolone, procarbizine, streptozotocin,tenoposide, 6-thioguanine, thioTEPA, topotecan, vinblastine,vincristine, vinorelbine, VP-16 and VM-26. In some embodiments the kitcomprises Cyclophospharnide, Doxorubicin, and Prednisone (CHP). In someembodiments the kit comprises Cyclophosphamide, Doxorubicin andPrednisolone. In some embodiments the kit comprises doxorubicin,vinblastine, and dacarbazine (AVD).

In another embodiment, provided herein is an article of manufacture orkit comprising the formulations described herein for administration inan auto-injector device. An auto-injector can be described as aninjection device that upon activation, will deliver its contents withoutadditional necessary action from the patient or administrator. They areparticularly suited for self-medication of therapeutic formulations whenthe delivery rate must be constant and the time of delivery is greaterthan a few moments.

It is understood that the aspects and embodiments described herein arefor illustrative purposes only and that various modifications or changesin light thereof will be suggested to persons skilled in the art and areto be included within the spirit and purview of this application andscope of the appended claims.

The invention will be more fully understood by reference to thefollowing examples. They should not, however, be construed as limitingthe scope of the invention. It is understood that the examples andembodiments described herein are for illustrative purposes only and thatvarious modifications or changes in light thereof will be suggested topersons skilled in the art and are to be included within the spirit andpurview of this application and scope of the appended claims.

EXAMPLES Example 1. Anti-CD30 Antibody Drug-Conjugate ImpairsProliferating Inducible T Regulatory Cells In Vitro

CD4⁺ T cells, isolated from healthy donor PBMC (Astarte Biologics,Bothell Wash.) were used to generate inducible T regulatory cells(iTregs). iTreg differentiation was performed over 1-2 weeks in 6-welltissue culture plates at 37° C. Cells were incubated with CD3/CD28 MACSiBead particles (Miltenyi biotec) at a 1:32 bead/cell ratio, in 2-3 mlof X-VIVO 15 media (Lonza) containing IL-2 (50 ng/ml), TGFβ (50 ng/ml),and a 1:100 dilution of Lipid-Mixture 1 (Sigma-Aldrich). iTregs wereevaluated for FoxP3 and CD30 expression by FACS analysis on an AttuneNXT flow cytometer (Life Technologies). Following differentiation,individual donor iTreg populations ranged between 20-80% FoxP3⁺ and40-70% CD30⁺.

To evaluate the effect of brentuximab vedotin (BV), and anti-CD30antibody drug-conjugate on iTreg viability, cells were driven toproliferate in vitro in the presence of BV or control antibodydrug-conjugate (IgG-MMAE). Briefly, iTregs were mixed with CD3/CD28beads (8:1) in RPMI 10% FCS and were distributed at approximately2.0×10⁴ cells/well into a 96-well round-bottom plate. A titration of BVor control IgG-MMAE was added to replicate wells at the concentrationslisted, for a final volume of 200 μl, and plates were incubated at 37°C. for 4-5 days. On the final day of the assay cells were stained withZombie Aqua Viability Dye and a non-competing monoclonal αCD30-PEantibody (Biolegend) for FACS analysis. BV drove a dose-dependentreduction in total viable iTreg numbers from five separate donors (FIG.1A). As populations of differentiated iTregs displayed heterogeneousCD30 expression, and BV selectively targets CD30 expressing cells,numbers of CD30⁺ iTregs were determined. Consistent with the loss oftotal iTregs from culture, BV showed enhanced depletion of CD30⁺ iTregs(FIG. 1B).

Example 2. Treatment with an Anti-CD30 Antibody Drug-Conjugate ReducesiTreg Numbers, De-Repressing In Vitro CD8⁺ T Cell Proliferation

Addition of T regulatory cells to activated CD8⁺ T cells in vitrofunctionally suppresses CD8⁺ T cell proliferation. iTregs, generated asdescribed above from two separate donors, showed suppressive activity onproliferating autologous CD8⁺ T cells in vitro. As shown in FIG. 2A andFIG. 2B, increasing the iTreg/CD8⁺ T cell ratio, for each donor, furtherabrogated T cell expansion, confirming suppressive function. To evaluatethe effect of BV on iTreg suppression in vitro, co-culture suppressionassays were performed. iTregs and CD8⁺ T cells were mixed at a 1:2 ratioand combined with CD3/CD28 beads. Replicate wells were treated with atitration of BV or control IgG-MMAE. After four days of culture, viableiTregs and CD8⁺ T cells were quantified by flow cytometry. As shown inFIG. 2C and FIG. 2D, increasing concentrations of BV selectively reducediTregs, resulting in augmented CD8⁺ T cell accumulation for two separatedonors.

Example 3. An Anti-CD30 Antibody Drug-Conjugate DepletesNaturally-Occurring Peripheral Blood Tregs, but not CD8⁺ T Cells InVitro

CD25^(hi) CD127^(lo) T regulatory cells or CD8⁺ T cells, enriched fromperipheral blood derived leukoreduction system (LRS) chambers, wereplated with CD3/CD28 beads+IL-2 in round-bottom 96-well tissue cultureplates for 4-5 days with a titration of BV or control IgG-MMAE. On thefinal day of the assay, cells were stained as described above (viabilitydye and αCD30-PE antibody) and evaluated by flow cytometry. For eachdonor, >50% of activated peripheral blood Tregs and CD8⁺ T cellsexpressed CD30, demonstrating abundant antigen expression for BVtargeting. As with iTreg cultures, BV drove a dose-dependent reductionof viable CD30⁺ Treg numbers from four separate donors (FIG. 3A). Incontrast, up to 10% g/ml of BV did not deplete CD30⁺ CD8⁺ T cells (FIG.3B).

Example 4. An Anti-CD30 Antibody Drug-Conjugate Reduces Human TRegulatory Cells and Increases CD8/Treg Ratio in a Xeno-GVHD Model

To evaluate activity of BV on activated human T cell subtypes in vivo, amodel of acute xenograft-driven graft-versus-host disease (xeno-GVHD)was employed. In this model, immune deficient NSG mice are lightlyirradiated (2Gy) on day 0 followed by adoptive transfer of 5×10⁶ healthydonor PBMC on day 1. Disease course is driven by activation andproliferation of mouse-reactive human CD4⁺ and CD8⁺ T cells, and diseasekinetics are slowed by addition of human T regulatory cells.

To evaluate the effect of BV on activated CD8⁺ T cells and Tregs invivo, Xeno-GVHD mice received a single i.p. injection of PBS or BV (3mg/kg) in PBS on day 5. On day 12, spleens were harvested and manuallydissociated through a 70% m cell strainer. Following centrifugation,individual spleens were resuspended in 3 ml of ACK lysis buffer (Sigma)for 3 minutes to remove red blood cells. Cells were washed with RPMI+10%FCS to stop the RBC lysis reaction. Spleen cells were resuspended in 4mls of media and 200 μl of the cell suspension was used for staining andanalysis by flow cytometry (FACS). Spleen cell suspensions were stainedwith Zombie Aqua Viability Dye (Biolegend) followed by staining withfluorescently labeled antibodies targeting human CD3, CD8, CD4, FoxP3,CD25, CD45, and murine CD45.1 (1:50 dilution, Biolegend) in stainingbuffer (PBS, 2% FCS, 1% NRS, 0.05% NaN₃) at 4° C. for 30 minutes. Cellswere washed and resuspended in 120%1 of staining buffer for plate-basedFACS using an Attune NXT flow cytometer. All events were collected from80%1 of sample, and FACS-measured cell concentrations were used tocalculate numbers of human immune cells. CD8⁺ T cells were identified asviability dye^(neg), hCD45⁺, mCD45.1⁻, CD3⁺, CD8⁺ cells. Tregs wereidentified as viability dye^(neg), hCD45⁺, mCD45.1⁻, CD3⁺, CD4⁺, FoxP3⁺,CD25⁺ cells.

As shown in FIG. 4A, BV significantly reduced human T regulatory cellsin the spleen compared to PBS alone. In contrast, splenic CD8⁺ T cellswere unaffected by BV treatment with a trend toward increased numbers(FIG. 4B). Taken together, BV treatment increased the CD8⁺ T cell/Tregratio in vivo consistent with heightened cytotoxic T cell activity (FIG.4C).

Example 5. An Anti-CD30 Antibody Drug-Conjugate Reduced CD30⁺ TRegulatory Cells in Patients with Classical Hodgkin Lymphoma

The effects of BV on circulating immune cells has not previously beenfully elucidated. Sixty-two patients adult patients with classicalHodgkin lymphoma (cHL) that had relapsed or was refractory to frontlinechemotherapy were enrolled in a study to evaluate treatment with BV.Patients were excluded if they previously received prior salvagetherapy, including salvage radiotherapy, for refractory cHL; BV or anyimmuno-oncology therapy affecting the PD-1, CTLA4, or CD137 pathways;and/or allogeneic or autologous stem cell transplant (ASCT). BV wasadministered to the patients at a dose of 1.8 mg/kg on Day 1 and thepatients were assessed on Day 8. Immunophenotyping of peripheral bloodby flow cytometry was performed by Q2 Solutions on heparinized wholeblood. Cell pellets, resulting from plasma banking, were sent toAdaptive for T Cell Receptor β (TCRβ) sequencing. Peripheral bloodmononuclear cells were isolated from CPT tubes, frozen, and thenanalyzed in batches by Caprion using their intracellular cytokinestaining platform following peptide stimulation.

BV treatment appears resulted in the reduction of T helper cell subsetpopulations including regulatory T cells (FIG. 5 ). As BV is aCD30-targeted therapeutic, and as CD30 is expressed transiently onimmune cells including B and T cells, CD30 expression on peripheralblood cells in the patients was evaluated. Regulatory T cells expressedmore CD30 than any other T cell subset examined (FIG. 6 ). TheseCD30-expressing regulatory T cells were significantly reduced in numberin the peripheral blood after treatment with BV (FIG. 7 ). In addition,BV treatment resulted in the reduction of CD30⁺ regulatory T cells moresignificantly when compared to the reduction of other CD30⁺ T helpercells.

Example 6. CD30 Expression is Enriched on CD25^(hi) CCR4^(hi) FOXP3^(hi)Effector Tregs in PBMC

Cryopreserved PBMCs from healthy donors were stained with viability dye,anti-CD3, CD4, CD8, CD45RA, FoxP3, CCR4, CD127 and CD25 (1:50 dilution,Biolegend) and evaluated by flow cytometry. Memory and naïve T cellpopulations were discriminated by CD45RA expression. T regulatory cellswere identified by appropriate expression of CD4, CD25, FoxP3 and/orCD127. As shown in FIG. 8A, CD30 is most frequently expressed by Tregulatory cells compared to CD4⁺ and CD8⁺ memory and naïve T cellsubsets. Furthermore, subdividing FoxP3-expressing Tregs into CD25^(hi)and CD25^(low/neg) populations showed expression of CD30 is highlyassociated with the effector T regulatory subset (FoxP3^(hi) CD25^(hi)CCR4^(hi)) (FIG. 8B).

Example 7. Differences in T Cell Subset CD30 Expression and Drug EffluxMay Underlie Sensitivity to BV

T cell subsets (CD4⁺, CD8⁺, CD4⁺ CD25⁺ CD127⁻) were sorted fromcryopreserved PBMC by magnetic selection and activated with CD3/CD28beads (1:4) in vitro for 7 days. Each day, CD30 expression was monitoredby flow cytometry. Values from a representative donor are shown as theproportion of cells expressing CD30 (FIG. 9A) and the relative magnitudeof expression by Mean Fluorescence Intensity (MFI) (FIG. 9B). Enriched Tregulatory cells displayed heightened CD30 expression kinetics andoverall CD30 receptor levels compared to CD4⁺ and CD8⁺ T cells followingactivation.

To examine whether heightened CD30 expression on T regulatory cellstranslated into enhanced payload delivery, an internalization assay wasperformed. On day 4 of in vitro stimulation, at peak receptorexpression, T cell subsets were incubated with a conditionallyfluorescent anti-CD30 mAb for 6 hours. Along the incubation time-course,cells were analyzed by flow cytometry for intracellular payload releasevia activation of a quenched-fluorescent reporter-CD30 mAb construct(FIG. 9C). T regulatory cells showed accelerated and increased releaseof fluorescent payload relative to CD4⁺ and CD8⁺ T cells, consistentwith heightened CD30 expression at day 4. These data support theconclusion that differences in CD30 expression may facilitate enhanceddrug delivery to T regulatory cells (Tregs).

Sensitivity of cells to many chemotherapies, including MMAE, isinfluenced by cell-intrinsic drug efflux activity. T cell subsets wereevaluated for relative efflux pump activity using a rhodamine 123 effluxassay, following manufacturer's protocol (Chemicon International,Multidrug Resistance Direct Dye Efflux Assay). Enriched T cellpopulations were loaded with rhodamine 123, incubated in a 37° C. waterbath, and were measured for loss of fluorescence over a 5-hourtime-course by flow cytometry. T regulatory cells showed the slowestrhodamine-123 efflux among T cell subsets while CD8⁺ T cells showedrapid clearance of intracellular rhodamine-123 (FIG. 9D).

Altogether, activated T regulatory cells demonstrate heightened CD30receptor expression and payload delivery, along with impaired drugefflux capacity, providing mechanistic rationale for the observedsensitivity to BV relative to cytotoxic CD8⁺ T cells.

1. A method of decreasing the activity of CD30⁺ T regulatory (Treg)cells in a subject comprising administering to the subject anantibody-drug conjugate, wherein the antibody-drug conjugate comprisesan anti-CD30 antibody or an antigen-binding portion thereof conjugatedto a monomethyl auristatin.
 2. The method of claim 1, wherein decreasingthe activity of CD30⁺ Treg cells comprises a decrease in the number ofCD30⁺ Treg cells.
 3. The method of claim 2, wherein the number of CD30⁺Treg cells is decreased relative to the number of one or more othertypes of CD4⁺ T cells.
 4. The method of claim 3, wherein the one or moreother types of CD4⁺ T cells comprise Th1 cells, Th2 cells or Th17 cells.5. The method of claim 4, wherein the one or more other types of CD4⁺ Tcells comprise Th1 CD30⁺ cells, Th2 CD30⁺ cells or Th17 CD30⁺ cells. 6.The method of claim 5, wherein the number of CD30⁺ Treg cells isdecreased relative to the number of CD30⁺ Treg cells in the subjectprior to administration of the antibody-drug conjugate.
 7. The method ofclaim 1, wherein decreasing the activity of CD30⁺ Treg cells comprises adecrease in the function of CD30⁺ Treg cells.
 8. The method of claim 7,wherein the decrease in the function of CD30⁺ Treg cells is relative tothe function of CD30⁺ Treg cells in a subject prior to administration ofthe antibody-drug conjugate.
 9. A method of increasing the ratio of CD8⁺T cells to CD30⁺ T regulatory (Treg) cells in a subject having cancercomprising administering to the subject an antibody-drug conjugate,wherein the antibody-drug conjugate comprises an anti-CD30 antibody oran antigen-binding portion thereof conjugated to a monomethylauristatin.
 10. A method of modulating the immune response in a subjecthaving cancer comprising administering to the subject an antibody-drugconjugate, wherein the antibody-drug conjugate comprises an anti-CD30antibody or an antigen-binding portion thereof conjugated to amonomethyl auristatin, wherein the modulation comprises increasing theratio of CD8⁺ T cells to CD30⁺ T regulatory (Treg) cells in the subject.11. The method of claim 9, wherein the ratio of CD8⁺ T cells to CD30⁺Treg cells is increased relative to the ratio of CD8⁺ T cells to CD30⁺Treg cells in the subject prior to the administration of theantibody-drug conjugate.
 12. The method of claim 1, wherein the CD30⁺Treg cells are CD30⁺ inducible T regulatory (iTreg) cells or CD30⁺peripheral T regulatory (pTreg) cells.
 13. The method of claim 1,wherein the monomethyl auristatin is monomethyl auristatin E (MMAE). 14.The method of claim 1, wherein the monomethyl auristatin is monomethylauristatin F (MMAF). 15-16. (canceled)
 17. The method of claim 1,wherein the anti-CD30 antibody of the antibody-drug conjugate comprisesa heavy chain variable region and a light chain variable region, whereinthe heavy chain variable region comprises: (i) a CDR-H1 comprising theamino acid sequence of SEQ ID NO: 1; (ii) a CDR-H2 comprising the aminoacid sequence of SEQ ID NO: 2; and (iii) a CDR-H3 comprising the aminoacid sequence of SEQ ID NO: 3; and wherein the light chain variableregion comprises: (i) a CDR-L1 comprising the amino acid sequence of SEQID NO: 4; (ii) a CDR-L2 comprising the amino acid sequence of SEQ ID NO:5; and (iii) a CDR-L3 comprising the amino acid sequence of SEQ ID NO:6.
 18. The method of claim 17, wherein the anti-CD30 antibody of theantibody-drug conjugate comprises a heavy chain variable regioncomprising the amino acid sequence of SEQ ID NO: 7 and a light chainvariable region comprising the amino acid sequence of SEQ ID NO: 8.19-43. (canceled)
 44. The method of claim 1, wherein the method furthercomprises administering one or more additional therapeutic agentscapable of modulating the immune response.
 45. The method of claim 44,wherein the one or more additional therapeutic agents is not an antibodyor antigen-binding fragment thereof.
 46. The method of claim 44, whereinthe one or more additional therapeutic agents is an antibody orantigen-binding fragment thereof.
 47. The method of claim 1, wherein themethod further comprises administering one or more additionaltherapeutic agents. 48-62. (canceled)